\title{Jalview 2.5: A manual and introductory tutorial }
-\author{David Martin, James Procter, Andrew Waterhouse and Geoff Barton}
+\author{David Martin, James Procter, Andrew Waterhouse, Saif Shehata, Nancy Giang,Suzanne Duce and Geoff Barton}
\date{Manual version 1.2.3 6th May 2011}
\newcommand{\clearemptydoublepage}{\newpage{\pagestyle{empty}}\cleardoublepage}
{\Huge
-Jalview 2.8.2
-}
+Jalview 2.9.0b2}
\vspace{0.5in}
{\huge
-A Manual with Introductory Tutorial }
+Manual and Introductory Tutorial }
\vspace{2.4in}
{\large
David Martin, James Procter, Andrew Waterhouse, Saif Shehata, Nancy Giang,
+
+
Suzanne Duce and Geoff Barton
\vspace{2in}
-Manual Version 1.5
+Manual Version 1.6
% post CLS lifesci course on 15th January
% draft. Remaining items are AACon, RNA visualization/editing and Protein disorder analysis exercises.
-4th December 2014
+9th June 2016
\end{center}
\pagenumbering{roman}
\setcounter{page}{1}
\tableofcontents
-\clearemptydoublepage
+%\clearemptydoublepage
% \listoffigures
% \newpage
% \listoftables
-% \newpage
+\newpage
\pagenumbering{arabic}
\setcounter{page}{1}
-
\chapter{Basics}
\label{jalviewbasics}
\section{Introduction}
\subsection{Jalview}
Jalview is a multiple sequence alignment viewer, editor and analysis tool.
Jalview is designed to be platform independent (running on Mac, MS Windows, Linux
-and any other platforms that supports Java). Jalview is capable of editing and
+and any other platforms that support Java). Jalview is capable of editing and
analysing large alignments (thousands of sequences) with minimal degradation in
performance, and able to show multiple integrated views of the alignment and
other data. Jalview can read and write many common sequence formats including
FASTA, Clustal, MSF(GCG) and PIR.
-There are two types of Jalview program. The {\bf Jalview Desktop} is a stand
-alone application that provides powerful editing, visualization, annotation and
+There are two types of Jalview program. The {\bf Jalview Desktop} is a standalone
+application that provides powerful editing, visualization, annotation and
analysis capabilities. The {\bf JalviewLite} applet has the same core
visualization, editing and analysis capabilities as the desktop, without the
desktop's webservice and figure generation capabilities. It is designed to be
embedded in a web page,\footnote{A demonstration version of Jalview (Jalview Micro
Edition) also runs on a mobile phone but the functionality is limited to sequence
-colouring.} and includes a javascript API to allow customisable display of alignments for web sites such as
-{\bf Pfam}.\footnote{\url{http://pfam.sanger.ac.uk}}
+colouring.} and includes a javascript API to allow customisable display of
+alignments for web sites such as Pfam.\footnote{\url{http://pfam.xfam.org}}
-Jalview 2.8.2 was released in December 2014. The Jalview Desktop in this version
+The Jalview Desktop in this version
provides access to protein and nucleic acid sequence, alignment and structure
databases, and includes the Jmol\footnote{ Provided under the LGPL licence at
\url{http://www.jmol.org}} viewer for molecular structures, and the VARNA\footnote{Provided under GPL licence at \url{http://varna.lri.fr}} program for the visualization of RNA secondary structure. A
visualization of sequence alignments, and their interactive analysis. Tree
building, principal components analysis, physico-chemical property conservation
and sequence consensus analyses are built into the program. Web services enable
-Jalview to access on line alignment and secondary structure prediction programs,
+Jalview to access online alignment and secondary structure prediction programs,
as well as to retrieve protein and nucleic acid sequences, alignments, protein structures and sequence annotation. Sequences, alignments, trees, structures, features and alignment annotation may also be exchanged with the local filesystem. Multiple visualizations of an alignment may be worked on simultaneously, and the user interface provides a comprehensive set of controls for colouring and layout. Alignment views are dynamically linked with Jmol structure displays, a tree viewer and spatial cluster display, facilitating interactive exploration of the alignment's structure. The application provides its own Jalview project file format in order to store the current state of an alignment and analysis windows. Jalview also provides WYSIWIG\footnote{WYSIWIG: What You See Is What You Get.} style figure generation capabilities for the preparation of alignments for publication.
\begin{figure}[htbp]
\begin{center}
between Jalview, TOPALi and AstexViewer.} in 2004, enabling Andrew Waterhouse and
Jim Procter to re-engineer the original program to introduce contemporary developments
in bioinformatics and take advantage of the latest web and Java technology.
-Jalview's development was supported from 2009
-by an award from the BBSRC's Tools and Resources fund. Jalview's development is
-currently being funded by a BBSRC `Extending The Jalview Resource for
-Biological Sequence Alignment and Analysis' grant and a Wellcome Trust
-`Extending The Jalview Resource for Biological Sequence Alignment and Analysis'
-grant. In 2010, 2011, and 2012, Jalview benefitted from the
+Jalview's development has been supported from 2009
+onwards by BBSRC funding, and since 2014 by a
+Wellcome Trust Biomedical Resource grant\footnote{Wellcome grant number 101651/Z/13/Z}. In 2010, 2011, and 2012, Jalview benefitted from the
\href{http://code.google.com/soc/}{Google Summer of Code}, when Lauren Lui and Jan Engelhardt introduced new features for handling RNA alignments and secondary structure annotation, in collaboration with Yann Ponty.\footnote{\url{http://www.lix.polytechnique.fr/~ponty/}}
This tutorial is written in a manual format with short exercises where
appropriate, typically at the end of each section. This chapter concerns the
basic operation of Jalview and should be sufficient for those who want to
-load Jalview (Section \ref{startingjv}), open an alignment (Section \ref{loadingseqs}), perform basic editing and colouring (Section \ref{selectingandediting} and Section \ref{colours}), and produce publication
-and presentation quality graphical output (Section \ref{layoutandoutput}).
-
-Chapter \ref{analysisannotation} covers the additional visualization and
-analysis techniques that Jalview provides. This includes working with the
-embedded Jmol molecular structure viewer, building and viewing trees and PCA
+launch Jalview (Section \ref{startingjv}), open an alignment (Section
+\ref{loadingseqs}), perform basic editing (Section
+\ref{selectingandediting}), colouring (Section \ref{colours}), and produce
+publication and presentation quality graphical output (Section \ref{layoutandoutput}).
+
+In addition, the manual covers the additional visualization and
+analysis techniques available in Jalview. This includes working
+with the embedded Jmol molecular structure viewer, building and viewing trees and PCA
plots, and using trees for sequence conservation analysis. An overview of
the Jalview Desktop's webservices is given in Section \ref{jvwebservices}, and
the alignment and secondary structure prediction services are described
-in detail in Sections \ref{msaservices} and \ref{protsspredservices}. Following
-this, Section \ref{featannot} details the creation and visualization of sequence
+in detail in Sections \ref{msaservices} and \ref{protsspredservices}. Section \ref{featannot} details the creation and visualization of sequence
and alignment annotation, and the retrieval of sequences and annotation from
-databases and DAS Servers. Finally, Section \ref{workingwithnuc} discusses
+databases and DAS Servers. Section \ref{workingwithnuc} discusses
specific features of use when working with nucleic acid sequences, such as translation and linking to protein
coding regions, and the display and analysis of RNA secondary structure.
enclosing the pressed keys with square brackets ({\em e.g.} [RETURN] or [CTRL]).
Keystroke combinations are combined with a `-' symbol ({\em e.g.} [CTRL]-C means
-press [CTRL] and the `C' key).
+press [CTRL] and the `C' key) simultaneously.
Menu options are given as a path from the menu
that contains them - for example {\sl File $\Rightarrow$ Input Alignment
$\Rightarrow$ From URL} means to select the `From URL' option from the `Input
Alignment' submenu of a window's `File' dropdown menu.
-\section{Obtaining and Starting the Jalview Desktop Application}
+\section{Launching the Jalview Desktop Application}
\label{startingjv}
\begin{figure}[htbp]
\begin{center}
The Jalview Desktop can be run in two ways; as an application launched from the
web {\sl via} Java webstart, or as an application loaded onto your hard drive.
The webstart version is launched from the pink `Launch Jalview Desktop
-button' at the top right handside of pages of the website
+button' at the top right hand side of pages of the website
\href{http://www.jalview.org}{(www.jalview.org)}.
To download the locally installable version, follow the links on the download
page
(Figure \ref{download}).
These links will launch the latest stable release of Jalview.\par
+%% this paragraph needs to be rewritten for the new signed certificate from Certum.
+
When the application is launched with webstart, two dialogs may appear before
the application starts. If your browser is not set up to handle webstart, then
clicking the launch link may download a file that needs to be opened
-manually, or prompt you to select the correct program to handle the webstart
+manually, or prompt you to select the program to handle the webstart
file. If that is the case, then you will need to locate the {\bf javaws} program
on your system\footnote{The file that is downloaded will have a type of {\bf
application/x-java-jnlp-file} or {\bf .jnlp}. The {\bf javaws} program that can run
\subsubsection{Jalview News RSS Feed}
Announcements are made available to users of the
-Jalview Desktop {\sl via} the Jalview News reader. This window will open
+Jalview Desktop {\sl via} the Jalview Newsreader. This window will open
automatically when new news is available, and can also be accessed {\sl via} the
Desktop's {\sl Tools $\Rightarrow$ Show Jalview News} menu entry.
\begin{figure}[htbp]
\begin{center}
\includegraphics[height=3in]{images/jvrssnews.pdf}
-\caption{{\bf The Jalview News Reader.} The news reader opens automatically
+\caption{{\bf The Jalview News Reader.} The newsreader opens automatically
when new articles are available from the Jalview Desktop's news channel.}
\label{jalviewrssnews}
\end{center}
\end{figure}
-\exercise{Launching Jalview from the Jalview website}{
+\exercise{Launching Jalview from the Jalview Website}{
\label{start}
\exstep{Open the Jalview web
site \href{http://www.jalview.org}{(www.jalview.org)}
in your web browser. Launch Jalview by clicking on the
pink `Launch Jalview' Desktop button in the top right hand corner. This
-will download and open a jalview.jnlp webstart file.} \exstep {Dialogue boxes
-will open and ask if you want to open the the jalview.jnlp file as the file is an application downloaded from the
+will download and open a jalview.jnlp webstart file.}
+\exstep {Dialogue boxes
+will open and ask if you want to open the jalview.jnlp file as the file is an application downloaded from the
Internet, click Open. (Note you maybe asked to update Java, if you agree then it
will automatically update the Java software). As Jalview opens, four demo
-Jalview windows automatically load.} \exstep {If
+Jalview windows automatically load.}
+\exstep {If
you are having trouble, it may help changing the browser you are using, as the browsers and
-it's version may effect this process.}
+it's version may affect this process.}
\exstep{To deactivate the opening of the 4 demo sequences during the launch, go
to the {\sl Tools $\Rightarrow$
Preferences...} menu on the desktop. A `Preference'
\exstep{To reload the original demo file select the
{\em File$\Rightarrow$ From URL} entry in the Desktop menu. Click on
the URL history button on the right hand side of the dialog box to view the
-files, selected exampleFile-2-7.jar, then click OK.}
+files, select exampleFile\_2\_7.jar, then click OK.}
{\bf Note:} Should you want to reload the example alignment or load your own
sequence during the launch process, then go
to the {\sl Tools $\Rightarrow$
Preferences...} menu on the desktop. The tick the `Open file' entry of `Visual'
preferences tab, type in the URL of the sequence you want to load. This
-file will load during the start up process.\\
-
+file will load during the start up process.
As the jalview.jnlp file launches Jalview on your desktop, you
may want to move this from the downloads folder to another folder.
Opening from this file will allow Jalview to be launched offline.
-{\bf Help launching Jalview is avaliable in a video on
-\href{http://www.jalview.org/Help/Getting-Started}{the Getting Started page} of
-the website.}}
+{\bf Help launching Jalview is available in videos on the Getting Started page
+of the Jalview website at \url{http://www.jalview.org/Help/Getting-Started}.}}
\subsection{Getting Help}
\label{gettinghelp}
-\subsubsection{Built in documentation}
+\subsubsection{Built in Documentation}
Jalview has comprehensive on-line help documentation. Select {\sl Help
$\Rightarrow$ Documentation} from the main window menu and a new window will
open (Figure \ref{help}). The appropriate topic can then be selected from the
\end{center}
\end{figure}
-\subsubsection{Email lists}
+\subsubsection{Email Lists}
The Jalview Discussion list {\tt jalview-discuss@jalview.org} provides a forum
for Jalview users and developers to raise problems and exchange ideas - any
editing and navigation is performed using the mouse, and {\bf cursor mode}
where editing and navigation are performed using the keyboard. The {\bf F2 key}
is used to switch between these two modes. With a Mac as the F2 is
- often assigned to screen brightness, one maay often need to type {\bf function
- [Fn] key with F2}.
+ often assigned to screen brightness, one may often need to type {\bf function
+ [Fn] key with F2} function
+ [Fn]-F2.
\begin{figure}[htb]
\begin{center}
\includegraphics[width=6.5in]{images/jalview_anatomy.pdf}
-\caption{{\bf The anatomy of Jalview.} The major features of the Jalview Desktop Application are labelled.}
+\caption{{\bf The anatomy of Jalview.} The major features of the Jalview Desktop Application are labeled.}
% TODO: modify text labels to be clearer - black on grey and black border for clarity
\label{anatomy}
\end{center}
\end{figure}
-\subsection{Navigation in Normal mode}
+\subsection{Navigation in Normal Mode}
Jalview always starts up in Normal mode, where the mouse is used to interact
with the displayed alignment view. You can move about the alignment by clicking
$\Rightarrow$ Close All} option from the Jalview desktop.
{\bf \em{Warning: make sure you have saved your work because this cannot be
-undone !}} }
+undone!}} }
\parbox[c]{3in}{\centerline{\includegraphics[width=2.5in]{images/start_closeall.pdf}
}}
-\subsection{Navigation in Cursor mode}
+\subsection{Navigation in Cursor Mode}
\label{cursormode}
\parbox[c]{5in}{Cursor mode navigation enables the user to quickly
and precisely navigate, select and edit parts of an alignment. On pressing F2 to
Rapid movement to specific positions is accomplished as listed below:
\begin{list}{$\circ$}{}
-\item {\bf Jump to Sequence {\sl n}:} Type a number {\sl n} then press [S] to move to sequence (row) {\sl n}
+\item {\bf Jump to Sequence {\sl n}:} Type a number {\sl n} then press [S] to
+move to sequence (row). {\sl n}
\item {\bf Jump to Column {\sl n}:} Type a number {\sl n} then press [C] to move to column {\sl n} in the alignment.
\item {\bf Jump to Residue {\sl n}:} Type a number {\sl n} then press [P] to move to residue number {\sl n} in the current sequence.
\item {\bf Jump to column {\sl m} row {\sl n}:} Type the column number {\sl m}, a comma, the row number {\sl n} and press [RETURN].
\end{list}
+\subsection{The Find Dialog Box}
+\label{searchfunction}
+A further option for navigation is to use the {\sl Select $\Rightarrow$
+Find\ldots} function. This opens a dialog box into which can be entered regular
+expressions for searching sequences and sequence IDs, or sequence numbers.
+Hitting the [Find next] button will highlight the first (or next) occurrence of
+that pattern in the sequence ID panel or the alignment, and will adjust the view
+in order to display the highlighted region. The Jalview Help provides
+comprehensive documentation for this function, and a quick guide to the regular
+expressions that can be used with it.
+%TODO insert a figure for the Find dialog box
\exercise{Navigation}{
-\label{navigate}
Jalview has two navigation and editing modes: {\bf normal} mode (where editing
and navigation are via the mouse) and the {\bf cursor} mode (where editing and
navigation are via the keyboard).
\exstep{Find the Overview Window, {\sl Views
$\Rightarrow$ Overview Window} and open it. Move around the
alignment by clicking and dragging the red box in the overview window.}
-\exstep{Look at the status bar (lower left hand corner of the alignment window) as
-you move the mouse over the alignment. It indicates information about the
+\exstep{Return to the alignment window. Look at the status bar (lower left hand
+corner of the alignment window) as you move the mouse over the alignment. It indicates information about the
sequence and residue under the cursor.}
-\exstep{Press [F2] to enter {\bf Cursor mode}. Use the direction {\bfarrow
-keys} to move the cursor around the alignment.}
+\exstep{Press [F2] key (or [Fn]/[F2] on Mac) to enter {\bf Cursor mode}. Use
+the direction keys to move the cursor around the alignment.}
\exstep{Move to sequence 7 by pressing {\bf 7 S}. Move to column 18 by pressing
{\bf 1 8 C}. Move to residue 18 by pressing {\bf 1 8 P}. Note that these can be
two different positions if gaps are inserted into the sequence. Move to sequence 5,
{\bf Note:} To view Jalview's comprehensive on-line help documentations select
Help on desktop menu, clicking on Documentation will open a Documentation
window. Select topic from the navigation panel on the left hand side or use the
-Search tab to select specific key words
-
-{\bf Help navigating Jalview is avaliable in a video on
-\href{http://www.jalview.org/Help/Getting-Started}{the Getting Started page} of
-the website.}}
-
-\subsection{The Find Dialog Box}
-\label{searchfunction}
-A further option for navigation is to use the {\sl Select $\Rightarrow$
-Find\ldots} function. This opens a dialog box into which can be entered regular
-expressions for searching sequences and sequence IDs, or sequence numbers.
-Hitting the [Find next] button will highlight the first (or next) occurrence of
-that pattern in the sequence ID panel or the alignment, and will adjust the view
-in order to display the highlighted region. The Jalview Help provides
-comprehensive documentation for this function, and a quick guide to the regular
-expressions that can be used with it.
-
-%TODO insert a figure for the Find dialog box
-
+Search tab to select specific key words.
+{\bf Help navigating is available in videos on the Getting Started page
+of the Jalview website at \url{http://www.jalview.org/Help/Getting-Started}}
+}
-\section{Loading your Own Sequences}
+\section{Loading Sequences and Alignments}
\label{loadingseqs}
-Jalview provides many ways to load your own sequences. %For this section of the tutorial you will need to download the file http://www.jalview.org/tutorial/alignment.fa to a suitable location on your hard drive.
-
+%Jalview provides many ways to load your own sequences. %For this section of the
+% tutorial you will need to download the file http://www.jalview.org/tutorial/alignment.fa to a suitable location on your hard drive.
\subsection{Drag and Drop}
In most operating systems you can just drag a file icon from a file browser
window and drop it on an open Jalview application window. The file will then be opened as a new alignment window. %You can try this with the tutorial file you have downloaded. When you have opened the file, close it again by selecting the close control on the window. If you drop an alignment file onto an open alignment window it will be appended.
\subsection{From a File}
Jalview can read sequence alignments from a sequence alignment file. This is a
-text file, not a word processor document. For entering sequences from a
+text file, {\bf not} a word processor document. For entering sequences from a
wordprocessor document see Cut and Paste (Section \ref{cutpaste}) below. Select
{\sl File $\Rightarrow$ Input Alignment $\Rightarrow$ From File} from the main
menu (Figure \ref{loadfile}). You will then get a file selection window where
registry. Jalview's sequence fetching capabilities allow you to avoid having to
manually locate and save sequences from a web page before loading them into
Jalview. It also allows Jalview to gather additional metadata provided by the
-source, such as annotation and database cross references.
+source, such as annotation and database cross-references.
Select {\sl File $\Rightarrow$ Fetch Sequence(s) \ldots} from the main menu and
a window will appear (Figure \ref{loadseq}). Pressing the database selection
button in the dialog box opens a new window showing all the database sources
\label{loadseq}
\end{center}
\end{figure}
-
+
+\subsection{Memory Limits}
+\label{memorylimits}
+Jalview is a Java program. One unfortunate implication of this is that Jalview
+cannot dynamically request additional memory from the operating system. It is
+important, therefore, that you ensure that you have allocated enough memory to
+work with your data. On most occasions, Jalview will warn you when you have
+tried to load an alignment that is too big to fit in to memory (for instance,
+some of the PFAM alignments are {\bf very} large). You can find out how much
+memory is available to Jalview with the desktop window's {\sl $\Rightarrow$
+Tools $\Rightarrow$ Show Memory Usage} function, which enables the display of
+the currently available memory at the bottom left hand side of the Desktop
+window's background. Should you need to increase the amount of memory available
+to Jalview, full instructions are given in the built in documentation (opened by
+selecting {\sl Help $\Rightarrow$ Documentation}) and on the JVM memory
+parameters page (\url{http://www.jalview.org/jvmmemoryparams.html}).
-\exercise{Loading sequences}{
+\exercise{Loading Sequences}{
\label{load}
\exstep{Use {\sl Window $\Rightarrow$ Close All} from the Desktop window menu to
close all windows.}
{\sl Window $\Rightarrow$ Close All} menu option from the Desktop.
Then type the same URL (\url{http://www.jalview.org/tutorial/alignment.fa}) into
your web browser and {\bf save} the file to your desktop.
-
Open the file you have just saved in Jalview by selecting {\sl File
$\Rightarrow$ Input Alignment $\Rightarrow$ From File} from the desktop menu and
selecting this file.
-Click OK to load the alignment.}
+Click OK to load.}
\exstep{{\bf Loading sequences by `Drag and Drop' / `Cut and Paste':}
(i) Select {\sl Desktop $\Rightarrow$ Window
drop it onto the Jalview window, the alignment should open.
Test the differences
-between (a) dragging the sequence onto the empty Jalview desktop and (b)
+between (a) dragging the sequence onto the Jalview desktop and (b)
dragging the sequence onto an existing alignment window.
(ii) Open \url{http://www.jalview.org/tutorial/alignment.fa} in a
-web browser. Drag the URL directly from browser window onto Jalview.
-
-(If the URL is downloaded instead of opened in the browser, then
-locate the file in your download directory and open it in a text editor.)
+web browser. Drag the URL directly from browser onto Jalview desktop. (If
+the URL is downloaded, then locate the file in your
+download directory and open it in a text editor.)
(iii) Open the alignment.fa file using text editor. Copy the sequence text from the
file into the clipboard and paste it into the desktop
\exstep{{\bf Loading sequences from Public Database:} Select {\sl File
$\Rightarrow$ Fetch Sequence(s)...} from the Desktop to open up new window
called New Sequence Fetcher.
-Press database selection button at the top of the dialog box, this opens
+Press database selection button (top of the dialog box), this opens
another window called Select Database Retrieval Source showing all the database
sources.
-Select the {\bf PFAM seed} database and click ok, then enter the accession
-number {\bf PF03460} and click OK. An alignment of about 107 sequences should load.
-
-Several database IDs
-or accession numbers can be loaded by using semicolons to separated them.}}
-
-\subsection{Memory Limits}
-\label{memorylimits}
-Jalview is a Java program. One unfortunate implication of this is that Jalview
-cannot dynamically request additional memory from the operating system. It is
-important, therefore, that you ensure that you have allocated enough memory to
-work with your data. On most occasions, Jalview will warn you when you have
-tried to load an alignment that is too big to fit in to memory (for instance,
-some of the PFAM alignments are {\bf very} large). You can find out how much
-memory is available to Jalview with the desktop window's {\sl $\Rightarrow$
-Tools $\Rightarrow$ Show Memory Usage} function, which enables the display of
-the currently available memory at the bottom left hand side of the Desktop
-window's background. Should you need to increase the amount of memory available
-to Jalview, full instructions are given in the built in documentation (opened by
-selecting {\sl Help $\Rightarrow$ Documentation}) and on the JVM memory
-parameters page (\url{http://www.jalview.org/jvmmemoryparams.html}).
-
+Select the {\bf PFAM seed} database and click OK, then enter the accession
+number {\bf PF03460} and click OK. An alignment of about 174 sequences should
+load. These can be viewed using the Overview window accessible from {\sl View
+$\Rightarrow$ Overview Window.}
+Several database IDs can be loaded by using semicolons to separate them.}
+{\bf Help loading sequences is available in videos on the Getting Started page
+of the Jalview website at \url{http://www.jalview.org/Help/Getting-Started}.}
+}
-\section{Writing Sequence Alignments}
-\label{savingalignments} \subsection{Saving the Alignment} Jalview allows the
+\section{Saving Sequences and Alignments}
+\label{savingalignments}
+\subsection{Saving Alignments} Jalview allows the
current sequence alignments to be saved to file so they can be restored at a
later date, passed to colleagues or analysed in other programs. From the
alignment window menu select {\sl File $\Rightarrow$ Save As} and a dialog box
PF03460 (see Exercise \ref{load}). } \exstep{
Select {\sl File $\Rightarrow$ Save As} from the alignment window menu. Choose a
-location into which to save the alignment and select your prefered format. All
+location into which to save the alignment and select your preferred format. All
formats except {\sl Jalview } jvp can be viewed in a normal text editor (e.g.
Notepad) or in a web browser.
Enter a file name and click {\sl Save}.
Close all windows and then load the project {\sl via} the {\sl File
$\Rightarrow$ Load Project} menu option. Observe how all the windows and
-positions are exactly as they were when they were saved. } }
+positions are exactly as they were when they were saved. }
+{\bf Help saving sequences is available in videos on the Getting Started page
+of the Jalview website at \url{http://www.jalview.org/Help/Getting-Started}.}}
-\section{Selecting and Editing Sequences}
+\chapter{Selecting and Editing Sequences }
+\label{jalviewediting}
+
\label{selectingandediting}
Jalview makes extensive use of selections - most of the commands available from
its menus operate on the {\sl currently selected region} of the alignment, either to
change their appearance or perform some kind of analysis. This section
illustrates how to make and use selections and groups.
-\subsection{Selecting Parts of an Alignment}
-Selections can be of arbitrary regions in an alignment, one or more complete columns, or one or more complete sequences.
-
-A selected region can be copied and pasted as a new alignment using the {\sl Edit $\Rightarrow$ Copy} and {\sl Edit $\Rightarrow$ Paste $\Rightarrow$ To New Alignment} in the alignment window menu options.
-
+\section{Selecting Parts of an Alignment}
+Selections can be of arbitrary regions in an alignment, one or more complete columns, or one or
+more complete sequences.
+A selected region can be copied and pasted as a new alignment
+using the {\sl Edit $\Rightarrow$ Copy} and {\sl Edit $\Rightarrow$ Paste $\Rightarrow$ To New
+Alignment} in the alignment window menu options.
{\bf To clear (unselect) the selection press the [ESC] (escape) key.}
-\subsubsection{Selecting Arbitrary Regions}
+\subsection{Selecting Arbitrary Regions}
To select part of an alignment, place the mouse at the top left corner of the region you wish to select. Press and hold the mouse button and drag the mouse to the bottom right corner of the chosen region then release the mouse button. A dashed red box appears around the selected region (Figure \ref{select}).
%[fig 12]
\end{center}
\end{figure}
-\subsubsection{Selecting Columns}
+\subsection{Selecting Columns}
To select the same residues in all sequences, click and drag along the alignment ruler.
This selects the entire column of the alignment. Ranges of positions from the
alignment ruler can also be selected by clicking on the first position and then
\end{center}
\end{figure}
-\subsubsection{Selecting Sequences}
+\subsection{Selecting Sequences}
\begin{figure}[htb]
\begin{center}
ranges of sequences (Figure \ref{selectrows}).
%[fig 14]
-\subsubsection{Making Selections in Cursor Mode}
+\subsection{Making Selections in Cursor Mode}
To define a selection in cursor mode (which is enabled by pressing [F2] when the alignment window is selected),
navigate to the top left corner of the proposed selection (using the mouse, the arrow keys, or the keystroke
commands described in Section \ref{cursormode}). Pressing the [Q] key marks this as the
-corner. A red outline appears around the cursor (Figure \ref{cselect})
+corner. A red outline appears around the cursor (Figure \ref{cselect}).
Navigate to the bottom right corner of the proposed selection and press the [M] key. This marks the bottom right corner of the selection. The selection can then be treated in the same way as if it had been created in normal mode.
\includegraphics[width=1.5in]{images/csel3.pdf}
\includegraphics[width=1.5in]{images/csel4.pdf}
\caption{{\bf Making a selection in cursor mode.} Navigate to the top left
-corner (left), press [Q] (left center), navigate to the bottom right corner (right center) and press [M] (right).}
+corner (left), press [Q], navigate to the bottom right corner and press [M] (right).}
\label{cselect}
\end{center}
\end{figure}
-\subsubsection{Inverting the Current Selection}
+\begin{figure}
+\begin{center}
+\includegraphics[width=3in]{images/group1.pdf}
+\includegraphics[width=1.5in]{images/newgroup.pdf}
+\caption{{\bf Creating a new group from a selection.}}
+\label{makegroup}
+\end{center}
+\end{figure}
+\subsection{Inverting the Current Selection}
The current sequence or column selection can be inverted, using {\sl Select
$\Rightarrow$ Invert Sequence/Column Selection} in the alignment window.
Inverting the selection is useful when selecting large regions in an alignment,
simply select the region that is to be kept unselected, and then invert the selection.
-This may also be useful when hiding large regions in an alignment (see Section \ref{hidingregions} below).
-Instead of selecting the columns and rows that are to be hidden, simply select the region that is to be kept
+This may also be useful when hiding large regions in an alignment (see Section \ref{hidingregions}
+below).
+Instead of selecting the columns and rows that are to be hidden, simply select the
+region that is to be kept
visible, invert the selection, then select {\sl View $\Rightarrow$ Hide
-$\Rightarrow$ Selected Region }.
+$\Rightarrow$ Selected Region}.
-\subsection{Creating Groups}
+\section{Creating Groups}
Selections are lost as soon as a different region is selected. Groups can be
created which are labeled regions of the alignment. To create a group, first
select the region which is to comprise the group. Then click the right mouse
`Group', `Edit Group Name', or `JGroupXXXXX' (Where XXXXX was some serial
number).} then enter a name for the group in the dialogue box which appears.
-\begin{figure}
-\begin{center}
-\includegraphics[width=3in]{images/group1.pdf}
-\includegraphics[width=1.5in]{images/newgroup.pdf}
-\caption{{\bf Creating a new group from a selection.}}
-\label{makegroup}
-\end{center}
-\end{figure}
-
By default the new group will have a box drawn around it. The appearance of the group can be changed (see Section \ref{colours} below). This group will stay defined even when the selection is removed.
-\subsection{Exporting the Current Selection}
-
-The current selection can be copied to the clipboard (in PFAM format). It can
-also be output to a textbox using the output functions in the pop-up menu
-obtained by right clicking the current selection. The textbox enables quick
-manual editing of the alignment prior to importing it into a new window (using
-the [New Window] button) or saving to a file with the {\sl File $\Rightarrow$
-Save As } pulldown menu option from the text box.
-
-
\exercise{Making Selections and Groups}{
\label{exselect}
-\exstep{Close all windows in Jalview.
-Load the ferredoxin alignment (PFAM
-ID PF03460 from PFAM seed database).
+\exstep{Close windows.
+Load the ferredoxin alignment (PF03460 from PFAM seed database).
Choose a residue and place the mouse
-cursor on it. Note residue information will show in alignment window status bar.
-Click and drag the mouse cursor to create a selection. As you drag, a red box
+cursor on it (Residue information will show in alignment window status
+bar)
+Click and drag the mouse to create a selection. As you drag, a red box
will `rubber band' out to
show the extent of the selection.
Release the mouse
-button and a red box should border the selected region.
-Press [ESC] to clear the selection.}
+button and a red box borders the selected region.
+Press [ESC] to clear this.}
\exstep{ Select one sequence by clicking on
the sequence ID panel. Note that the sequence ID takes on a highlighted
background and a red box appears around the selected sequence.
Hold down [SHIFT] and click another sequence ID a few positions above or below.
Note how the selection expands to include all the sequences between the two positions on which you clicked.
-
Hold down [CTRL] and then click on several sequences ID's both selected and
unselected. Note how unselected IDs are individually added to the selection and previously selected IDs are
individually deselected.}
Press {\bf Q} to mark this position.
Navigate to column 65, row 8 by pressing {\bf 6 5 , 8 [RETURN]}. Press {\bf M}
to complete the selection. Note to clear the selection press the {\bf[ESC]}
-key.} \exstep{To create a {\bf group} from the selected the region, click the
-right mouse button when mouse is on the selection, this opens the selection
-popup menu in the alignment window.
+key.}
+\exstep{To create a {\bf group} from the selected the region, click the
+right mouse button when mouse is on the selection, this opens a
+pop-up menu in the alignment window.
Open the {\sl Selection $\Rightarrow$ Edit New Group $\Rightarrow$ Group Colour
} menu and select `Percentage Identity'.
This will turn the selected region into a group and colour it accordingly.}
\exstep{Hold down [CTRL] and use the mouse to select and deselect sequences in
the alignment by clicking on their Sequence ID label. Note how the group expands
-to include newly selected sequences, and the `Percentage Identity' colouring changes. } \exstep{ Use the mouse to click and drag the right-hand edge of the selected group. Note again how the group resizes.}
+to include newly selected sequences, and the `Percentage Identity' colouring changes. }
+\exstep{ Another way to resize the group is by using the mouse to click and drag
+the right-hand edge of the selected group.}
\exstep{The current selection can be {\bf exported} and saved by right clicking
-on the text area to open the selection popup-menu. Follow the menus and pick an
+on the text area to open the Sequence ID pop-up menu. Follow the menus and pick an
output format (eg BLC) from the {\sl Selection $\Rightarrow$ Output to Textbox
\ldots} submenu.
}
\exstep{In the alignment output window, try manually editing the alignment,
importing group into a new alignment window by clicking the [New Window] button to import the
file into a new alignment window.}
-
+{\bf Additional help is available from videos on the Jalview website at \url{http://www.jalview.org/training/Training-Videos}.}
% more? change colouring style. set border colour.
}
-\subsection{Reordering the Alignment}
+\section{Exporting the Current Selection}
+The current selection can be copied to the clipboard (in PFAM format). It can
+also be output to a textbox using the output functions in the pop-up menu
+obtained by right clicking the current selection. The textbox enables quick
+manual editing of the alignment prior to importing it into a new window (using
+the [New Window] button) or saving to a file with the {\sl File $\Rightarrow$
+Save As } pulldown menu option from the text box.
+
+\section{Reordering an Alignment}
Sequence reordering is simple. Highlight the sequences to be moved then press the up or down arrow keys as appropriate (Figure \ref{reorder}). If you wish to move a sequence up past several other sequences it is often quicker to select the group past which you want to move it and then move the group rather than the individual sequence.
\begin{figure}[htbp]
\exstep{To select and move multiple
sequences, use hold [SHIFT] and [CTRL], and select two sequences separated by
one or more un-selected sequences. Note how multiple sequences are grouped
-together when they are re-ordered using the up and down arrow keys.} }
+together when they are re-ordered using the up and down arrow keys.}
+{\bf Additional help is available from videos on the Jalview website
+at \url{http://www.jalview.org/training/Training-Videos}.}}
-\subsection{Hiding Regions}
+\section{Hiding Regions}
\label{hidingregions}
It is sometimes convenient to exclude some sequences or residues in the alignment without actually deleting them. Jalview allows sequences or alignment columns within a view to be hidden, and this facility has been used to create the several different views in the example alignment file that is loaded when Jalview is first started (See Figure \ref{startpage}).
$\Rightarrow$ All but Selected Region } menu entry, or press [Shift]+[Ctrl]+H
to hide the unselected region.
-\subsubsection{Representing a Group with a Single Sequence}
+\subsection{Representing a Group with a Single Sequence}
Instead of hiding a group completely, it is sometimes useful to work with just one representative sequence. The {\sl $<$Sequence ID$>$ $\Rightarrow$ Represent group with $<$Sequence ID$>$ } option from the sequence ID pop-up menu enables this variant of the hidden groups function. The remaining representative sequence can be visualized and manipulated like any other. However, any alignment edits that affect the sequence will also affect the whole sequence group.
-\exercise{Hiding and revealing regions}{
+\exercise{Hiding and Revealing Regions}{
\exstep{Close all windows, open the PFAM accession PF03460. Select a
contiguous set of sequences by clicking and dragging on the sequence ID panel.
-Right click on the selected sequence IDs to bring up the selection pop-up menu, select {\sl Hide Sequences}.
+Right click on the selected sequence IDs to bring up the sequence ID pop-up
+menu, select {\sl Hide Sequences}.
}
\exstep{
Right click on the blue triangle indicating hidden sequences and select {\sl
Reveal Sequences} in the panel. (If you have hidden all sequences then you will
-need to use the alignment window menu option {\sl View $\Rightarrow$ Show $\Rightarrow$ All Sequences.}) }
+need to use the alignment window menu option {\sl View $\Rightarrow$ Show $\Rightarrow$
+All Sequences.}) }
\exstep{
Repeat the process but use a non-contiguous set of sequences. Note that when
multiple regions are hidden there are two options, {\sl Reveal Sequences} and {\sl Reveal All}.
}
-\exstep{Repeat the above using columns to hide and reveale columns
-instead of sequences.
-}
+\exstep{Repeat the above using columns to hide and reveal columns
+instead of sequences.}
\exstep{Select a region of the alignment, then add in some additional columns to
the selection, and experiment with the `Hide all but selected region' function
in {\sl View $\Rightarrow$ Hide $\Rightarrow$ All but selected region.}}
-\exstep{Select some sequences and pick one to represent the rest by hoovering
-the mouse over this sequence. Bring up the selection pop-up menu by right
-clicking and then seelct {\sl (Sequence ID) $\Rightarrow$ Represent group
-with (Sequence ID)}. To reveal these hidden sequences, right click on the
-Sequence ID and in the the pop-up menu select Reveal All.}}
-
+\exstep{Select some sequences and pick one to represent the rest by hovering
+the mouse over this sequence. Bring up the Sequence ID pop-up menu by right
+clicking and then select {\sl (Sequence ID name) $\Rightarrow$ Represent group
+with (Sequence ID name )}. To reveal these hidden sequences, right click on the
+Sequence ID and in the pop-up menu select Reveal All.}
+{\bf Additional help is available from videos on the Jalview
+website at \url{http://www.jalview.org/training/Training-Videos}}}
+\begin{figure}[htb]
+\begin{center}
+\includegraphics[width=3in]{images/edit1.pdf}
+\includegraphics[width=3in]{images/edit2.pdf}
+\caption{{\bf Introducing gaps in a single sequence.} Gaps are introduced as the
+selected sequence is dragged to the right while pressing and holding [SHIFT].}
+\label{gapseq}
+\end{center}
+\end{figure}
-\subsection{Introducing and Removing Gaps}
+\begin{figure}[htb]
+\begin{center}
+\includegraphics[width=3in]{images/edit3.pdf}
+\includegraphics[width=3in]{images/edit4.pdf}
+\caption{{\bf Introducing gaps in a group.} Gaps are introduced as the selected
+group is dragged to the right with [CTRL] pressed.}
+\label{gapgroup}
+\end{center}
+\end{figure}
+\section{Introducing and Removing Gaps}
The alignment view provides an interactive editing interface, allowing gaps to be inserted or deleted to the left of any position in a sequence or sequence group. Alignment editing can only be performed whilst in keyboard editing mode (entered by pressing [F2]) or by clicking and dragging residues with the mouse when [SHIFT] or [CTRL] is held down (which differs from earlier versions of Jalview).
-\subsubsection{Locked Editing}
+\subsection{Locked Editing}
\label{lockededits}
The Jalview alignment editing model is different to that used in other alignment
-editors. Because edits are restricted to the insertion and deletion of gaps to the left of a particular sequence position, editing has the effect of shifting the rest of the sequence(s) being edited down or up-stream with respect to the rest of alignment. The {\sl Edit $\Rightarrow$ Pad Gaps} option can be enabled to eliminate `ragged edges' at the end of the alignment, but does not avoid the `knock-on' effect which is sometimes undesirable. However, its effect can be limited by performing the edit within a selected region. In this case, gaps will only be removed or inserted within the selected region. Edits are similarly constrained when they occur adjacent to a hidden column.
-
-\subsubsection{Introducing gaps in a single sequence}
-
-To introduce a gap, place the cursor on the residue to the immediate right of where the gap should appear. Hold down the SHIFT key and the left mouse button, then drag the sequence to the right till the required number of gaps has been inserted.
+editors. Because edits are restricted to the insertion and deletion of gaps to the left of a particular sequence position,
+editing has the effect of shifting the rest of the sequence(s) being edited down or up-stream with respect to the rest of
+alignment. The {\sl Edit $\Rightarrow$ Pad Gaps} option can be enabled to eliminate `ragged edges' at the end of the alignment,
+but does not avoid the `knock-on' effect which is sometimes undesirable.
+However, its effect can be limited by performing the edit within a selected region.
+In this case, gaps will only be removed or inserted within the selected region.
+Edits are similarly constrained when they occur adjacent to a hidden column.
+
+\subsection{Introducing Gaps in a Single Sequence}
+To introduce a gap, first select the sequence in the sequence ID panel and
+then place the cursor on the residue to the immediate right of where the gap
+should appear. Hold down the SHIFT key and the left mouse button, then drag the sequence to the right until the required number of gaps has been inserted.
One common error is to forget to hold down [SHIFT]. This results in a selection which is one sequence high and one residue long. Gaps cannot be inserted in such a selection. The selection can be cleared and editing enabled by pressing the [ESC] key.
-\subsubsection{Introducing gaps in all sequences of a group}
-
-To insert gaps in all sequences in a selection or group, place the mouse cursor on any residue in the selection or group to the immediate right of the position in which a gap should appear. Hold down the CTRL key and the left mouse button, then drag the sequences to the right until the required number of gaps has appeared.
+\subsection{Introducing Gaps in all Sequences of a Group}
+To insert gaps in all sequences in a selection or group, select the
+required sequences in the sequence ID panel and then place the mouse cursor on
+any residue in the selection or group to the immediate right of the position in which a gap should appear. Hold down the CTRL key and the left mouse button, then drag the sequences to the right until the required number of gaps has appeared.
Gaps can be removed by dragging the residue to the immediate right of the gap
leftwards whilst holding down [SHIFT] (for single sequences) or [CTRL] (for a group of sequences).
-\subsubsection{Sliding Sequences}
+\subsection{Sliding Sequences}
Pressing the [$\leftarrow$] or [$\rightarrow$] arrow keys when one or more
-sequences are selected will ``slide'' the selected sequences to the left or
-right (respectively). Slides occur regardless of the region selection -
+sequences are selected will ``slide'' the entire selected sequences to the left
+or right (respectively). Slides occur regardless of the region selection -
which, for example, allows you to easily reposition misaligned subfamilies
within a larger alignment.
-
-\subsubsection{Undoing edits}
-Jalview supports the undoing of edits {\sl via} the {\sl Edit $\Rightarrow$ Undo Edit}
-alignment window menu option, or CTRL-Z. An edit, if undone, may be re-applied
-with {\sl Edit $\Rightarrow$ Redo Edit}, or CTRL-Y. Note, however, that the
-{\sl Undo} function only works for edits to the alignment or sequence ordering.
-Colouring of the alignment, showing and hiding of sequences or modification of
-annotation cannot be undone.
-
-
% % better idea to introduce hiding sequences, and use the invert selection, hide
% others, to simplify manual alignment construction
-\exercise{Editing alignments}{
-\label{mousealedit}
+
+\exercise{Editing Alignments}
+ %\label{mousealedit}
% TODO: VERIFY FOR 2.6.1 and 2.7 - NUMBERING/INSTRUCTIONS APPEAR OFF
-{You are going to manually reconstruct part of the example Jalview alignment
- available at
- \href{http://www.jalview.org/examples/exampleFile.jar}{http://www.jalview.org/examples/exampleFile.jar}.
-}
- \item{Remember to use [CTRL]+Z to undo an edit, or the {\sl File $\Rightarrow$
+{You are going to manually reconstruct part of the example Jalview
+alignment available at
+ \href{http://www.jalview.org/examples/exampleFile.jar}
+ {http://www.jalview.org/examples/exampleFile.jar}.
+Remember to use [CTRL]+Z to undo an edit, or the {\sl File $\Rightarrow$
Reload } function to revert the alignment back to the original version if you
- want to start again.}
- \item{ If you are using OSX, and a key combination - such as [CTRL]+A - does
+ want to start again.
+If you are using OSX, and a key combination - such as [CTRL]+A - does
not work, then try pressing the [CMD] key instead of [CTRL].
- }
\exstep{ Load the URL
\textsf{http://www.jalview.org/tutorial/unaligned.fa} which contains part of the
-ferredoxin alignment from PF03460.
-}
+ferredoxin alignment from PF03460.}
\exstep{ Select the first 7 sequences, and press H to hide them (or right click
-on the sequence IDs to open the sequence ID popup menu, and select {\sl Hide
-Sequences}). }
+on the sequence IDs to open the sequence ID pop-up menu, and select {\sl Hide
+Sequences}).}
\exstep{ Select FER3\_RAPSA and FER\_BRANA. Slide the sequences to
-the left so the initial {\bf A} lies at column 57 using the $\Rightarrow$ key.}
+the right so the initial {\bf A} lies at column 57 using the $\Rightarrow$ key.}
\exstep{ Select FER1\_SPIOL, FER1\_ARATH, FER2\_ARATH, Q93Z60\_ARATH and
-O80429\_MAIZE (Hint: you can do this by pressing [CTRL]-I to invert the sequence selection and then
+O80429\_MAIZE
+(Hint: you can do this by pressing [CTRL]-I to invert the sequence selection and then
deselect FER1\_MAIZE), and use the $\Rightarrow$ key to slide them to so they
begin at column 5 of the alignment view.}
\exstep{ Select all the visible
-sequences in the block by pressing [CTRL]-A. Insert a single gap in all selected
-sequences at column 38 by holding [CTRL] and clicking on the R in FER1\_SPIOL and
-dragging one column to right. Insert another gap at column 47 in all sequences in
-the same way.}
+sequences (those not hidden) in the block by pressing [CTRL]-A.
+Insert a single
+gap in all selected sequences at column 38 of the alignment by holding [CTRL]
+and clicking on the R at column 38 in the FER1\_SPIOL, then drag one
+column to right.
+Insert another gap at column 47 in all sequences in the same way.}
\exstep{ Correct the ferredoxin domain alignment for FER1\_SPIOL by
inserting two additional gaps after the gap at column 47: First press [ESC] to
\exstep{ Now complete the
alignment of FER1\_SPIOL with a {\bf locked edit} by pressing [ESC] and select
columns 47 to 57 of the FER1\_SPIOL row. Move the mouse onto the G at column 50,
-hold [SHIFT] and drag the G to the left by one column to insert a gap at column
-57.}
+hold [SHIFT] and drag the G in column 47 of FER1\_SPIOL to the left by one
+column to insert a gap at column 57.}
-\exstep{ In the next two steps you will complete the alignment of the last two sequences.
+\exstep{ In the next two steps you will complete the alignment of the last two
+sequences.
Select the last two sequences (FER1\_MAIZE and O80429\_MAIZE), then press [SHIFT]
and click and drag the initial methionine of O80429\_MAIZE 5 columns to the right
-so it lies at column 10. Keep holding [SHIFT] and click and drag to insert
+so it lies at column 10.
+
+Keep holding [SHIFT] and click and drag to insert
another gap at the proline at column 25 (25C in cursor mode). Remove the gap at
column 44, and insert 4 gaps at column 47 (after AAPM).}
{\sl Edit $\Rightarrow$ Undo Edit} and {\sl Edit $\Rightarrow$ Redo Edit} menu
option, or their keyboard shortcuts ([CTRL]+Z and [CTRL]+Y) to step
backwards and replay the edits you have made.}
-
}
-\begin{figure}[htb]
-\begin{center}
-\includegraphics[width=3in]{images/edit1.pdf}
-\includegraphics[width=3in]{images/edit2.pdf}
-\caption{{\bf Introducing gaps in a single sequence.} Gaps are introduced as the
-selected sequence is dragged to the right while pressing and holding [SHIFT].}
-\label{gapseq}
-\end{center}
-\end{figure}
-
-\begin{figure}[htb]
-\begin{center}
-\includegraphics[width=3in]{images/edit3.pdf}
-\includegraphics[width=3in]{images/edit4.pdf}
-\caption{{\bf Introducing gaps in a group.} Gaps are introduced as the selected
-group is dragged to the right with [CTRL] pressed.}
-\label{gapgroup}
-\end{center}
-\end{figure}
-
-
-\subsubsection{Editing in Cursor mode}
-
-Gaps can be be easily inserted when in cursor mode (toggled with [F2]) by
+\subsection{Editing in Cursor mode}
+Gaps can be easily inserted when in cursor mode (toggled with [F2]) by
pressing [SPACE]. Gaps will be inserted at the cursor, shifting the residue
under the cursor to the right. To insert {\sl n} gaps type {\sl n} and then
press [SPACE]. To insert gaps into all sequences of a group, use [CTRL]-[SPACE]
columns in all sequences in the selected group, and those columns are to the
right of the selected residue.
-\exercise{Keyboard edits}{ \item{This continues on from exercise
+\section{Undoing Edits}
+Jalview supports the undoing of edits {\sl via} the {\sl Edit $\Rightarrow$ Undo Edit}
+alignment window menu option, or CTRL-Z. An edit, if undone, may be re-applied
+with {\sl Edit $\Rightarrow$ Redo Edit}, or CTRL-Y. Note, however, that the
+{\sl Undo} function only works for edits to the alignment or sequence ordering.
+Colouring of the alignment, showing and hiding of sequences or modification of
+annotation cannot be undone.
+
+\exercise{Keyboard Edits}
+{This continues on from exercise
\ref{mousealedit}, and recreates the final part of the example ferredoxin
alignment from the unaligned sequences using Jalview's keyboard editing mode.
-{\bf {\sl Note for Windows Users:}} The [SHIFT]-[SPACE] command has the same effect as
-the [CTRL]-[SPACE] command mentioned in this exercise, and you should use
-[SHIFT]-[SPACE] in order to avoid opening the window menu.}
+{\bf Note:} For Mac users, [CTRL]-[SPACE] command
+has the same effect as the [SHIFT]-[SPACE] command mentioned in this exercise.
+Window users should use [SHIFT]-[SPACE] rather than the [CTRL]-[SPACE] command,
+as this command will close the window.
\exstep{Load the sequence alignment at
\textsf{http://www.jalview.org/tutorial/unaligned.fa}, or continue using the
edited alignment from exercise \ref{mousealedit}. If you continue from the
-previous exercise, then first right click on the sequence ID panel and select
-{\sl Reveal All}.
-
-Now, enter cursor mode by pressing [F2]}
+previous exercise, first right click on the sequence ID panel and select
+{\sl Reveal All}. Enter cursor mode by pressing [F2].}
% TODO: BACKSPACE or DELETE WHEN SEQS ARE SELECTED WILL DELETE ALL SEQS JAL-783
-\exstep{Insert 58 gaps at the start of the first sequence (FER\_CAPAA). Press {\sl 58} then {\sl [SPACE]}. }
-\exstep{Go down one sequence and select rows 2-5 as a block. Click on the second sequence ID (FER\_CAPAN). Hold down shift and click on the fifth (FER1\_PEA). }
-\exstep{Insert 6 gaps at the start of this group. Go to column 1 row 2 by typing {\sl 1,2} then pressing {\sl [RETURN]}. Now insert 6 gaps. Type {\sl 6} then hold down {\sl [CTRL]} and press {\sl [SPACE]}.}
+
+\exstep{Insert 58 gaps at the start of the sequence 1 (FER\_CAPAA). Press
+{\sl 58} then {\sl [SPACE]}. }
+
+\exstep{Go down one sequence and select rows 2-5 as a block. Click on the second sequence ID (FER\_CAPAN).
+ Hold down shift and click on the fifth (FER1\_PEA). }
+
+\exstep{Insert 6 gaps at the start of this group. Go to column 1 row 2 by typing
+{\sl 1,2} then press {\sl [RETURN]}. Now insert 6 gaps in all the sequences.
+Type {\sl 6} then hold down {\sl [SHIFT]} and press {\sl [SPACE]}.}
\exstep{Now insert one gap at column 34 and another at 38. Insert 3 gaps at 47.
-Press {\sl 34C} then {\sl [CTRL]-[SPACE]}. Press {\sl 38C} then [CTRL]-[SPACE].
-Press {\sl 47C} then {\sl 3 [CTRL-SPACE]} the first through fourth sequences are
-now aligned.}
+Press {\sl 34C} then {\sl [SHIFT]-[SPACE]}. Press {\sl 38C} then
+[SHIFT]-[SPACE].
+Press {\sl 47C} then {\sl 3 [SHIFT-SPACE]} the first through fourth sequences
+are now aligned.}
\exstep{The fifth sequence (FER1\_PEA) is poorly aligned. We will delete some gaps and add some new ones. Press {\sl [ESC]} to clear the selection. Navigate to the start of sequence 5 and delete 3 gaps. Press {\sl 1,5 [RETURN]} then {\sl 3 [BACKSPACE]} to delete three gaps. Go to column 31 and delete the gap. Press {\sl 31C [BACKSPACE]} .}
-\exstep{ Similarly delete the gap now at column 34, then insert two gaps at column 38. Press {\sl 34C [BACKSPACE] 38C 2 [SPACE]}. Delete three gaps at column 44 and insert one at column 47 by pressing {\sl 44C 3 [BACKSPACE] 47C [SPACE]}. The top five sequences are now aligned.}
+\exstep{ Similarly delete the gap now at column 34, then insert two gaps at
+column 38.
+Press {\sl 34C [BACKSPACE] 38C 2 [SPACE]}. Delete three gaps at column 44 and
+insert one at column 47 by pressing {\sl 44C 3 [BACKSPACE] 47C [SPACE]}. The top five sequences are
+now aligned.}
}
+\chapter{Colouring Sequences and Figure Generation}
+\label{colouringfigures}
\section{Colouring Sequences}
\label{colours}
displayed, and you may have to disable it using the {\sl View $\Rightarrow$
Show Features} option before you can see your colourscheme.
-There are two main types of colouring styles: simple static residue colourschemes and dynamic schemes which use conservation and consensus analysis to control colouring. A hybrid colouring is also possible, where static residue schemes are modified using a dynamic scheme. The individual schemes are described in Section \ref{colscheme} below.
+There are two main types of colouring styles: {\bf simple static residue}
+colourschemes and {\bf dynamic schemes} which use conservation and consensus
+analysis to control colouring. {\bf Hybrid colouring} is also possible, where
+static residue schemes are modified using a dynamic scheme. The individual schemes are described in Section \ref{colscheme} below.
\subsection{Colouring the Whole Alignment}
\subsection{Colouring a Group or Selection}
-Selections or groups can be coloured in two ways. The first is {\sl via} the Alignment Window's {\sl Colour} menu as stated above, after first ensuring that the {\sl Apply Colour To All Groups} flag is not selected. This must be turned {\sl off} specifically as it is {\sl on} by default. When unticked, selections from the Colours menu will only change the colour for residues in the current selection, or the alignment view's ''background colourscheme'' when no selection exists.
+Selections or groups can be coloured in two ways. The first is {\sl via} the Alignment Window's {\sl Colour} menu as stated above,
+ after first ensuring that the {\sl Apply Colour To All Groups} flag is {\bf
+ not} selected.
+ This must be turned {\sl off} specifically as it is {\sl on} by default.
+ When unticked, selections from the Colours menu will only change the colour for residues in the current selection,
+ or the alignment view's ``background colourscheme'' when no selection exists.
-The second method is to use the {\sl Selection $\Rightarrow$ Group $\Rightarrow$ Group Colour} context menu option obtained by right clicking on the group (Figure \ref{colgrp}). This only changes the colour of the current selection or group.
+The second method is to select sequences and right click mouse to open pop-up
+menu and select {\sl Selection $\Rightarrow$ Edit New Group $\Rightarrow$ Group
+Colour} from context menu options
+(Figure \ref{colgrp}). This only changes the colour of the current selection or group.
\begin{figure}[htbp]
\begin{center}
}\parbox[c]{3in}{
\centerline{\includegraphics[width=2.8in]{images/col_byannot.pdf}}}
-The {\bf per Sequency only} option in the {\bf Colour By Annotation} dialog
+The {\bf per Sequence} option in the {\bf Colour By Annotation} dialog
allows each sequence to be shaded according to sequence associated annotation
-rows, such as protein disorder scores. This functionality is described futher in
-Section \ref{protdisorderpred}.
+rows, such as protein disorder scores. This functionality is described further
+in Section \ref{protdisorderpred}.
\subsection{Colour Schemes}
\subsubsection{Taylor}
\parbox[c]{3.5in}{
-This colour scheme was devised by Willie Taylor and an entertaining description of it's origin can be found in Protein Engineering, Vol 10 , 743-746 (1997)
+This colour scheme was devised by Willie Taylor and an entertaining description of its origin can be found in Protein Engineering,
+Vol 10 , 743-746 (1997).
}
\parbox[c]{3in}{
\includegraphics[width=2.75in]{images/col_taylor.pdf}
\parbox[c]{3.5in}{ Residues are coloured according to whether the corresponding
nucleotide bases are purine (magenta) or pyrimidine (cyan) based. All non ACTG
residues are uncoloured. For further information about working with nucleic acid
-sequences and alignments, see Section \ref{workingwithnuc}
+sequences and alignments, see Section \ref{workingwithnuc}.
%and Section \ref{workingwithrna}
-.
} \parbox[c]{3in}{ \includegraphics[width=2.75in]{images/col_purpyr.pdf} }
-\subsubsection{RNA Helix colouring}
+\subsubsection{RNA Helix Colouring}
\parbox[c]{3.5in}{ Columns are coloured according to their assigned RNA helix as
defined by a secondary structure annotation line on the alignment. Colours for
each helix are randomly assigned, and option only available when an RNA
\includegraphics[width=2.75in]{images/col_rnahelix.pdf} }
\exercise{Colouring Alignments}{
+\exstep{Ensuring that the {\sl Apply Colour To All Groups} flag is not selected
+in View sequence alignment menu.
+ This must be turned {\sl off} specifically as it is {\sl on} by default.}
\exstep{
-Open a sequence alignment, for example the PFAM domain PF03460. Select the alignment menu option {\sl Colour $\Rightarrow$ ClustalX}. Note the colour change. Now try all the other colour schemes in the {\sl Colour} menu. Note that some colour schemes do not colour all residues.
+Open a sequence alignment, for example the PFAM domain PF03460 in PFAM seed.
+Select the alignment menu option {\sl Colour $\Rightarrow$ ClustalX}. Note the colour change. Now try all the other colour schemes in the {\sl Colour} menu. Note that some colour schemes do not colour all residues.
}
\exstep{
-Colour the alignment using {\sl Colour $\Rightarrow$ Blosum62}. Select a group of around 4 similar sequences. Use the context menu (right click on the group) option {\sl Selection $\Rightarrow$ Group $\Rightarrow$ Group Colour $\Rightarrow$ Blosum62} to colour the selection. Notice how some residues which were not coloured are now coloured. The calculations performed for dynamic colouring schemes like Blosum62 are based on the group being coloured, not the whole alignment (this also explains the colouring changes observed in exercise \ref{exselectgrpcolour} during the group selection step).
+Colour the alignment using {\sl Colour $\Rightarrow$ Blosum62}. Select a group
+of around 4 similar sequences. Use the context menu (right click on the group)
+option {\sl Selection $\Rightarrow$ Edit New Group $\Rightarrow$ Group Colour
+$\Rightarrow$ Blosum62} to colour the selection. Notice how some residues which
+were not coloured are now coloured. The calculations performed for dynamic
+colouring schemes like Blosum62 are based on the group being coloured, not the
+whole alignment (this also explains the colouring changes observed in exercise
+\ref{exselect} during the group selection step).
}
\exstep{
-Keeping the same selection as before, colour the complete alignment using {\sl Colour $\Rightarrow$ Taylor}. Select the menu option {\sl Colour $\Rightarrow$ By Conservation}. Slide the selector from side to side and observe the changes in the alignment colouring in the selection and in the complete alignment.
+Keeping the same selection as before, colour the complete alignment except
+the group using {\sl Colour $\Rightarrow$ Taylor}.
+Select the menu option {\sl Colour $\Rightarrow$ By Conservation}.
+Slide the selector from side to side and observe the changes in the alignment colouring in the selection and in the complete alignment.
}
}
\end{figure}
-\exercise{User defined colour schemes}{
+\exercise{User Defined Colour Schemes}{
\exstep{Load a sequence alignment. Select the alignment menu option {\sl Colour $\Rightarrow$ User Defined}. A dialogue window will open.
}
\exstep{Click on an amino acid button, then select a colour for that amino acid. Repeat till all amino acids are coloured to your liking.
}
}
-\section{Alignment Formatting and Graphics Output}
+\section{Formatting and Graphics Output}
\label{layoutandoutput}
-Jalview is a WYSIWIG alignment editor. This means that for most kinds of graphics output, the layout that is seen on screen will be the same as what is output in an exported graphics file. It is therefore important to pick the right kind of display layout prior to generating figures.
+Jalview is a WYSIWIG alignment editor. This means that for most kinds of graphics output,
+the layout that is seen on screen will be the same as what is outputted in an
+exported graphics file.
+It is therefore important to pick the right kind of display layout prior to generating figures.
\subsection{Multiple Alignment Views}
\subsection{Alignment Layout}
Jalview provides two screen layout modes, unwrapped (the default) where the alignment is in one long line across the window, and wrapped, where the alignment is on multiple lines, each the width of the window. Most layout options are controlled by the Format menu option in the alignment window, and control the overall look of the alignment in the view (rather than just a selected region).
-\subsubsection{Wrapped alignments}
+\subsubsection{Wrapped Alignments}
Wrapped alignments can be toggled on and off using the {\sl Format $\Rightarrow$ Wrap} menu option (Figure \ref{wrap}). Note that the annotation lines are also wrapped. Wrapped alignments are great for publications and presentations but are of limited use when working with large numbers of sequences.
If annotations are not all visible in wrapped mode, expand the alignment window to view them. Note that alignment annotation (see Section \ref{featannot}) cannot be interactively created or edited in wrapped mode, and selection of large regions is difficult.
\parbox[c]{4in}{The text appearance in a view can be modified {\sl via} the {\sl Format $\Rightarrow$ Font\ldots} alignment window menu. This setting applies for all alignment and annotation text except for that displayed in tool-tips. Additionally, font size and spacing can be adjusted rapidly by clicking the middle mouse button and dragging across the alignment window.}
\parbox[c]{2in}{\centerline{\includegraphics[width=1.75in]{images/font.pdf}}}
-\subsubsection{Numbering and label justification}
+\subsubsection{Numbering and Label Justification}
Options in the {\sl Format} menu are provided to control the alignment view, and provide a range of options to control the display of sequence and alignment numbering, the justification of sequence IDs and annotation row column labels on the annotation rows shown below the alignment.
-\subsubsection{Alignment and Group colouring and appearance}
+\subsubsection{Alignment and Group Colouring and Appearance}
The display of hidden row/column markers and gap characters can be turned off with {\sl Format $\Rightarrow$ Hidden Markers} and {\sl Format $\Rightarrow$ Show Gaps}, respectively. The {\sl Text} and {\sl Colour Text} option controls the display of sequence text and the application of alignment and group colouring to it. {\sl Boxes } controls the display of the background area behind each residue that is coloured by the applied coloursheme.
-\subsubsection{Highlighting nonconserved symbols}
+\subsubsection{Highlighting Nonconserved Symbols}
The alignment layout and group sub-menu both contain an option to hide
conserved symbols from the alignment display ({\sl Format $\Rightarrow$ Show
nonconserved } in the alignment window or {\sl Selection $\Rightarrow$ Group
The annotation lines which appear below the sequence alignment are described in
detail in Section \ref{featannot}. They can be hidden by toggling the {\sl View
$\Rightarrow$ Show Annotations} menu option. Additionally, each annotation line
-can be hidden and revealed in the same way as sequences {\sl via} the context menu on
-the annotation name panel (Figure \ref{annot}). Annotations can be reordered by
-dragging the annotation line label on the annotation label panel. Placing the
+can be hidden and revealed in the same way as sequences {\sl via} the
+pop-up context menu on the annotation name panel (Figure \ref{annot}).
+Annotations can be reordered by dragging the annotation line label on the annotation label panel. Placing the
mouse over the top annotation label brings up a resize icon on the left. When this is
displayed, Click-dragging up and down provides more space in the alignment window for viewing the annotations, and less space for the sequence alignment.
\exercise{Alignment Layout}{
\label{exscreen}
-\exstep{Start Jalview and open the URL \textsf{http://www.jalview.org/examples/exampleFile.jar}. Select {\sl Format $\Rightarrow$ Wrap} from the alignment window menu. Experiment with the various options from the {\sl Format} menu. to adjust the ruler placement, sequence ID format and so on. }
-\exstep{Hide all the annotation rows by selecting {\sl View $\Rightarrow$ Show Annotations} from the alignment window menu. Reveal the annotations by selecting the same menu option.}
-\exstep{{\bf Deselect} {\sl Format $\Rightarrow$ Wrap}, and right click on the annotation row labels to bring up the context menu. Select {\sl Hide This Row}. Bring up the context menu again and select {\sl Show All Hidden Rows} to reveal them}
+\exstep{Start Jalview and open the URL \textsf{http://www.jalview.org/examples/exampleFile.jar}.
+Select {\sl Format $\Rightarrow$ Wrap} from the alignment window menu.
+Experiment with the various options from the {\sl Format} menu, for example adjust the ruler placement,
+sequence ID format and so on. }
+\exstep{Hide all the annotation rows by selecting {\sl Annotations $\Rightarrow$
+Show Annotations} from the alignment window menu. Reveal the annotations by selecting the same menu option.} \exstep{Deselect {\sl Format $\Rightarrow$ Wrap}. Right click on the
+annotation row labels to bring up the pop-up context menu, then select {\sl
+Hide This Row}. Bring up the pop-up context menu again and select {\sl
+Show All Hidden Rows} to reveal them.}
\exstep{Annotations can be reordered by clicking and dragging the row to the desired position. Click on the {\sl Consensus} row and drag it upwards to just above {\sl Quality}. The rows should now be reordered. Features and annotations are covered in more detail in Section \ref{featannot} below.}
-\exstep{Move the mouse to the top left hand corner of the Secondary Structure annotation row label - a grey up/down arrow symbol should appear - when this is shown, the height of the {\sl Annotation Area} can be changed by Clicking and dragging the mouse up or down.}
+\exstep{Move the mouse to the top left hand corner of the Secondary Structure annotation row label -
+a grey up/down arrow symbol should appear - when this is shown, the height of the {\sl Annotation Area} can be changed
+by clicking and dragging the mouse up or down.}
}
\subsection{Graphical Output}
}
\parbox[c]{3.5in}{\centerline{\includegraphics[width=3in]{images/image_png.pdf}} \par \centerline{Zoom Detail of PNG image.}}
\exercise{Graphical Output}{
-\exstep{Load the example Jalview Jar file in Exercise \ref{exscreen}. Customise it how you wish but leave it unwrapped. Select {\sl File $\Rightarrow$ Export Image $\Rightarrow$ HTML} from the alignment menu. Save the file and open it in your favourite web browser. }
-\exstep{Now wrap the alignment and export the image to HTML again. Compare the two images. Note that the exported image matches the format displayed in the alignment window but {\bf annotations are not exported}.}
-\exstep{Export the alignment using the {\sl File $\Rightarrow$ Export Image $\Rightarrow$ PNG} menu option. Open the file in an image viewer that allows zooming such as Paint or Photoshop (Windows), or Preview (Mac OS X) and zoom in. Notice that the image is a bitmap and it becomes pixelated very quickly. Note also that the {\bf annotation lines are included} in the image.}
+\exstep{Load the example Jalview Jar file in Exercise \ref{exscreen}.
+Customise it how you wish but leave it unwrapped.
+Select {\sl File $\Rightarrow$ Export Image $\Rightarrow$ HTML} from the alignment menu.
+Save the file and open it in your favoured web browser. }
+\exstep{Wrap the alignment and export the image to HTML again. Compare the two
+images. (Note that the exported image matches the format displayed in the
+alignment window but {\bf annotations are not exported}).}
+\exstep{Export the alignment using the {\sl File $\Rightarrow$ Export Image $\Rightarrow$ PNG} menu option.
+Open the file in an image viewer that allows zooming such as Paint or Photoshop (Windows), or Preview (Mac OS X) and zoom in.
+Notice that the image is a bitmap and it becomes pixelated very quickly.
+(Note that the {\bf annotation lines are included} in the image.)}
\exstep{Export the alignment using the {\sl File $\Rightarrow$ Export Image
$\Rightarrow$ EPS} menu option. Open the file in a suitable program such as
Photoshop, Illustrator, Inkscape, Ghostview, Powerpoint (Windows), or
Preview (Mac OS X). Zoom in and note that the image has near-infinite
resolution.} }
-\chapter{Analysis and Annotation}
-\label{analysisannotation}
+\chapter{Annotation and Features}
+\label{featannot}
+Features and annotations are additional information that is overlaid on the sequences and the alignment. Generally speaking, annotations are associated with columns in the alignment. Features are associated with specific residues in the sequence.
-This chapter describes the annotation, analysis, and visualization tasks that
-the Jalview Desktop can perform.
+Annotations are shown below the alignment in the annotation panel, and often reflect properties of the alignment as a whole.
+Conversely, sequence features are properties of the individual sequences, so they do not change with the alignment,
+but are shown mapped on to specific residues within the alignment.
-Section \ref{wkwithstructure} introduces the structure visualization
-capabilities of Jalview. In Section \ref{alignanalysis}, you will find
-descriptions and exercises on building and displaying trees, PCA analysis,
-alignment redundancy removal, pairwise alignments and alignment conservation
-analysis. Section \ref{jvwebservices} introduces the various web based services
-available to Jalview users, and Section \ref{jabaservices} explains how to
-configure the Jalview Desktop for access to new JABAWS servers.
-Section \ref{msaservices} describes how to use the range of multiple alignment
-programs provided by JABAWS, and Section \ref{aacons} introduces JABAWS' AACon
-service for protein multiple alignment conservation analysis.
-Section \ref{protsspredservices} explains how to perform protein secondary
-structure predictions with JPred, and JABAWS' protein disorder prediction
-services are introduced in Section \ref{protdisorderpred}.
+Features and annotation can be interactively created, or retrieved from external
+data sources. DAS (the Distributed Annotation System) is the primary source of
+sequence features, whilst webservices like JNet (see \ref{jpred} above) can be used to analyse a
+given sequence or alignment and generate annotation for it.
+
+
+\section{Conservation, Quality and Conservation Annotation}
+\label{annotationintro}
+Jalview automatically calculates several quantitative alignment annotations
+which are displayed as histograms below the multiple sequence alignment columns.
+Conservation, quality and conservation scores are examples of dynamic
+annotation, so as the alignment changes, they change along with it.
+The scores can be used in the hybrid colouring options to shade the alignments.
+Mousing over a conservation histogram reveals a tooltip with more information.
+
+These annotations can be hidden and deleted via the context menu linked to the
+annotation row; but they are only created on loading an alignment. If they are
+deleted then the alignment should be saved and then reloaded to restore them.
+Jalview provides a toggle to autocalculate a consensus sequence upon editing. This is normally selected by default, but can be turned off for
+large alignments {\sl via} the {\sl Calculate $\Rightarrow$ Autocalculate
+Consensus} menu option if the interface is too slow.
-Section \ref{featannot} describes the mechanisms provided by Jalview for
-interactive creation of sequence and alignment annotation, and how they can be
-displayed, imported and exported and used to reorder the alignment. Section
-\ref{featuresfromdb} discusses the retrieval of database references and
-establishment of sequence coordinate systems for the retrieval and display of
-features from databases and DAS annotation services. Section
-\ref{workingwithnuc} describes functions and visualization techniques relevant
-to working with nucleotide sequences, coding region annotation and nucleotide
-sequence alignments.
-% and Section \ref{workingwithrna} covers the visualization,
-% editing and analysis of RNA secondary structure.
+\subsubsection{Conservation Annotation}
-\section{Working with structures}
-\label{wkwithstructure}
-Jalview facilitates the use of protein structures for the analysis of alignments
-by providing a linked view of structures associated with sequences in
-the alignment. The Java based molecular viewing program Jmol\footnote{See the
-Jmol homepage \url{http://www.jmol.org} for more information.} has been
-incorporated\footnote{Earlier versions of Jalview included MCView - a simple
-main chain structure viewer. Structures are visualized as an alpha carbon trace
-and can be viewed, rotated and coloured in a structure viewer and the results
-interpreted on a sequence alignment.} which enables sophisticated molecular
-visualizations to be prepared and investigated alongside an analysis of
-associated sequences.
-PDB format files can be imported directly or structures can be retrieved from
-the European Protein Databank (PDBe) using the Sequence Fetcher (see
-\ref{fetchseq}).
+Alignment conservation annotation is quantitative numerical index reflecting the
+conservation of the physico-chemical properties for each column of the alignment.
+The calculation is based on AMAS method of multiple sequence alignment analysis (Livingstone C.D. and Barton G.J. (1993) CABIOS Vol. 9 No. 6 p745-756),
+with identities scoring highest, and amino acids with substitutions in the same physico-chemical class have next highest score.
+The score for each column is shown below the histogram.
+The conserved columns with a score of 11 are indicated by '*'.
+Columns with a score of 10 have mutations but all properties are conserved are marked with a '+'.
-\subsection{Automatic association of PDB structures with sequences}
-Jalview can automatically determine which structures are associated with a
-sequence in a number of ways.
-\subsubsection{Discovery of PDB IDs from sequence database cross-references}
-If a sequence has an ID from a public database that contains cross-references to
-the PDB, such as Uniprot. Right-click on any sequence ID and select {\sl Structure $\Rightarrow$
-Associate Structure with Sequence $\Rightarrow$ Discover PDB IDs } from the context menu (Figure \ref{auto}). Jalview will attempt to associate the
-sequence with a Uniprot sequence and from there discover any associated PDB
-structures. This takes a few seconds and applies to all sequences in the
-alignment which have valid Uniprot IDs. On moving the cursor over the sequence
-ID the tool tip\footnote{Tip: The sequence ID tooltip can often become large for
-heavily cross referenced sequence IDs. Use the {\sl View $\Rightarrow$ Sequence
-ID Tooltip $\Rightarrow$ } submenu to disable the display of database cross
-references or non-positional features. } now shows the Uniprot ID and any
-associated PDB structures.
+\subsubsection{Consensus Annotation}
+
+Alignment consensus annotation reflects the percentage of the different residue
+per column. By default this calculation includes gaps in columns, gaps can be ignored via the Consensus label context
+menu to the left of the consensus bar chart.
+The consensus histogram can be overlaid
+with a sequence logo that reflects the symbol distribution at each column of
+the alignment. Right click on the Consensus annotation row and select the {\sl Show
+Logo} option to display the Consensus profile for the group or alignment.
+Sequence logos can be enabled by default for all new alignments {\sl via} the
+Visual tab in the Jalview desktop's preferences dialog box.
+
+\subsubsection{Quality Annotation}
+
+Alignment quality annotation is an ad-hoc measure of the likelihood of observing
+the mutations (if any) in a particular column of the alignment. The quality score is calculated for each column in an alignment by summing,
+for all mutations, the ratio of the two BLOSUM 62 scores for a mutation pair and each residue's conserved BLOSUM62 score (which is higher).
+This value is normalised for each column, and then plotted on a scale from 0 to 1.
+
+\subsubsection{Group Associated Annotation}
+\label{groupassocannotation}
+Group associated consensus and conservation annotation rows reflect the
+sequence variation within a particular group. Their calculation is enabled
+by selecting the {\sl Group Conservation} or {\sl Group Consensus} options in
+the {\sl Annotation $\Rightarrow$ Autocalculated Annotation } submenu of the
+alignment window.
+
+\subsection{Creating User Defined Annotation}
+
+Annotations are properties that apply to the alignment as a whole and are visualized on rows in the annotation panel.
+To create a new annotation row, right click on the annotation label panel and select the {\sl Add New Row} menu option (Figure \ref{newannotrow}).
+A dialogue box appears. Enter the label to use for this row and a new row will appear.
+
+To create a new annotation, first select all the positions to be annotated on the appropriate row.
+Right-clicking on this selection brings up the context menu which allows the insertion of graphics for secondary structure ({\sl Helix} or {\sl Sheet}),
+text {\sl Label} and the colour in which to present the annotation (Figure \ref{newannot}). On selecting {\sl Label} a dialogue box will appear,
+requesting the text to place at that position. After the text is entered, the selection can be removed and the annotation becomes clearly
+visible\footnote{When annotating a block of positions, the text can be partly obscured by the selection highlight. Pressing the [ESC] key clears
+the selection and the label is then visible.}. Annotations can be coloured or deleted as desired.
\begin{figure}[htbp]
\begin{center}
-%TODO fix formatting
-\parbox{1.5in}{
-{\centering
-\begin{center}
-\includegraphics[width=1.5in]{images/auto1.pdf}
-\end{center}}
-} \parbox{3.25in}{
-{\centering
-\begin{center}
-\includegraphics[scale=0.5]{images/auto2.pdf}
-\end{center}
-}
-} \parbox{1.5in}{
-{\centering
-\begin{center}
-\includegraphics[width=1.5in]{images/auto3.pdf}
-\end{center}
-}
-}
-
-\caption{{\bf Automatic PDB ID discovery.} The tooltip (left) indicates that no PDB structure has been associated with the sequence. After PDB ID discovery (center) the tool tip now indicates the Uniprot ID and any associated PDB structures (right)}
-\label{auto}
+\includegraphics[width=1.3in]{images/annots1.pdf}
+\includegraphics[width=2in]{images/annots2.pdf}
+\caption{{\bf Creating a new annotation row.} Annotation rows can be reordered by dragging them to the desired place.}
+\label{newannotrow}
\end{center}
\end{figure}
-\subsubsection{Drag-and-drop association of PDB files with sequences by filename
-match}
-\label{multipdbfileassoc}
-If one or more PDB files stored on your computer are dragged from their location
-on the file browser onto an alignment window, Jalview will search the alignment
-for sequences with IDs that match any of the files dropped onto the alignment.
-If it discovers matches, a dialog like the one in Figure
-\ref{multipdbfileassocfig} is shown, giving the option of creating associations
-for the matches.
-
-If no associations are made, then sequences extracted
-from the structure will be simply added to the alignment. However, if only
-some of the PDB files are associated, jalview will raise another dialog box giving
-you the option to add any remaining sequences from the PDB structure files not present in
-the alignment. This allows you to easily decorate sequences in a newly imported
-alignment with any corresponding structures you've already collected in a directory
-accessible from your computer.\footnote{We plan to extend this facility in
-future so Jalview will automatically search for PDB files matching your
-sequence within a local directory. Check out
-\href{http://issues.jalview.org/browse/JAL-801}{Jalview issue 801}}
-
-% there is no mention of the other footnote (#3) that appears saying: Tip: The sequence ID tooltip can often become large for heavily cross-referenced sequence IDs. Use the ...
-% JBP: yes there is - under 'Discovery of ' subsection.
\begin{figure}[htbp]
\begin{center}
-\includegraphics[]{images/pdbdragdropassoc.pdf}
-
-\caption{{\bf Associating PDB files with sequences by drag-and-drop.} Dragging
-PDB files onto an alignment of sequences with names matching the dragged files
-names (A), results in a dialog box (B) that gives the option to associate each
-file with any sequences with matching IDs. }
-\label{multipdbfileassocfig}
+\includegraphics[width=2in]{images/annots3.pdf}
+\includegraphics[width=2in]{images/annots4.pdf}
+\includegraphics[width=2in]{images/annots5.pdf}
+\caption{{\bf Creating a new annotation.} Annotations are created from a selection on the annotation row and can be coloured as desired.}
+\label{newannot}
\end{center}
\end{figure}
+\subsection{Automated Annotation of Alignments and Groups}
-\subsection{Viewing Structures}
-The structure viewer can be launched in two ways from the sequence ID context
-menu. To view a particular structure associated with a sequence in the
-alignment, simply select it from popup menu's associated structures submenu in
-{\sl Structure $\Rightarrow$ View Structure $\Rightarrow$ $<$PDB ID$>$}. The
-second way is most useful if you want to view all structural data available for
-a set of sequences in an alignment. If any of the {\bf currently selected}
-sequences have structures associated, the {\sl Structure } submenu of the
-sequence ID popup menu will include an option to {\sl View {\bf N}
-structures}. Selecting this option will open a new structure view containing
-the associated structures superposed according to the alignment.
+On loading a sequence alignment, Jalview will normally\footnote{Automatic
+annotation can be turned off in the {\sl Visual } tab in the {\sl Tools
+$\Rightarrow$ Preferences } dialog box.} calculate a set of automatic annotation
+rows which are shown below the alignment. For nucleotide sequence alignments,
+only an alignment consensus row will be shown, but for amino acid sequences,
+alignment quality (based on BLOSUM 62) and physicochemical conservation will
+also be shown. Conservation is calculated according to Livingstone and
+Barton\footnote{{\sl ``Protein Sequence Alignments: A Strategy for the
+Hierarchical Analysis of Residue Conservation." } Livingstone C.D. and Barton
+G.J. (1993) {\sl CABIOS } {\bf 9}, 745-756}.
+Consensus is the modal residue (or {\tt +} where there is an equal top residue).
+The inclusion of gaps in the consensus calculation can be toggled by
+right-clicking on the the Consensus label and selecting {\sl Ignore Gaps in
+Consensus} from the pop-up context menu located with consensus annotation row.
+Quality is a measure of the inverse likelihood of unfavourable mutations in the alignment. Further details on these
+calculations can be found in the on-line documentation.
-In both cases, each structure to be displayed will be downloaded or loaded from
-the local file system, and shown as a ribbon diagram coloured according to the
-associated sequence in the current alignment view (Figure \ref{structure}
-(right)). The structure can be rotated by clicking and dragging in the structure
-window. The structure can be zoomed using the mouse scroll wheel or by
-[SHIFT]-dragging the structure.
-Moving the mouse cursor over a sequence to which the structure is linked in the
-alignment view highlights the respective residue's sidechain atoms. The
-sidechain highlight may be obscured by other parts of the molecule. Similarly,
-moving the cursor over the structure shows a tooltip and highlights the
-corresponding residue in the alignment. Clicking the alpha carbon or phosphorous
-backbone atom will toggle the highlight and residue label on and off. Often, the
-position highlighted in the sequence may not be in the visible portion of the
-current alignment view and the sliders will scroll automatically to show the
-position. If the alignment window's {\sl View $\Rightarrow$ Automatic Scrolling
-} option is not selected, however, then the automatic adjustment will be
-disabled for the current view.
-\begin{figure}[htbp]
-\begin{center}
-\parbox{3in}{
-{\centering
-\begin{center}
-\includegraphics[scale=0.5]{images/structure1.pdf}
-\end{center}
-}
-}
-\parbox{3.2in}{
-{\centering
-\begin{center}
-\includegraphics[width=3in]{images/structure2.pdf}
-\end{center}
+\exercise{Annotating Alignments}{
+\exstep{Load the alignment at \textsf{http://www.jalview.org/tutorial/alignment.fa}.
+Right-click on the {\sl Conservation} annotation row to
+bring up the context menu and select {\sl Add New Row}. A dialogue box will appear asking for {\sl Label for annotation}.
+Enter ``Iron binding site" and click OK. A new, empty, row appears.
}
+\exstep{
+Navigate to column 97. Move down and on the new annotation row called
+``Iron binding site, select column 97.
+Right click at this selection and select {\sl Label} from the context menu.
+Enter ``Fe" in the box and click OK. Right-click on the selection again and select {\sl Colour}.
+Choose a colour from the colour chooser dialogue
+and click OK. Press [ESC] to remove the selection.
+}
+\exstep{ Select columns 70-77 on the annotation row. Right-click and choose {\sl Sheet} from the
+ context menu. You will be prompted for a label. Enter ``B" and press OK. A new line showing the
+ sheet as an arrow appears. The colour of the label can be changed but not the colour of the sheet
+ arrow.
+}
+\exstep{Right click on the title text of annotation row that you just created.
+Select {\sl Export Annotation} and, in the {\bf Export Annotation} dialog box that will open, select the Jalview format and click
+the [To Textbox] button.
+
+The format for this file is given in the Jalview help. Press [F1] to open it, and find
+the ``Annotations File Format'' entry in the ``Alignment Annotations'' section of the contents
+pane. }
+
+\exstep{Export the file to a text editor and edit the file to change the name of the annotation
+row. Save the file and drag it onto the alignment view.}
+\exstep{Try to add an additional helix somewhere along the row by editing the file and
+re-importing it.
+{\sl Hint: Use the {\bf Export Annotation} function to view what helix annotation looks like in
+a Jalview annotation file.}}
+\exstep{Use the {\sl Alignment Window $\Rightarrow$ File $\Rightarrow$ Export Annotations...}
+function to export all the alignment's annotation to a file.}
+\exstep{Open the exported annotation in a text editor, and use the {\bf Annotation File Format}
+documentation to modify the style of the Conservation, Consensus and Quality annotation rows so
+they appear as several lines on a single line graph.
+{\sl Hint: You need to change the style of annotation row in the first field of the annotation
+row entry in the file, and create an annotation row grouping to overlay the three quantitative
+annotation rows.}
}
-\caption{{\bf Structure visualization} The structure viewer is launched from the sequence ID context menu (left) and allows the structure to be visualized using the embedded Jmol molecular viewer (right). }
-\label{structure}
-\end{center}
-\end{figure}
+\label{viewannotfileex}\exstep{Recover or recreate the secondary structure
+prediction that you made in exercise \ref{secstrpredex}. Use the {\sl File $\Rightarrow$ Export
+Annotation} function to view the Jnet secondary structure prediction annotation row. Note the
+{\bf SEQUENCE\_REF} statements surrounding the row specifying the sequence association for the
+annotation. } }
-\subsection{Customising structure display}
-Structure display can be modified using the {\sl Colour} and {\sl View} menus
-in the structure viewer. The background colour can be modified by selecting the
-{\sl Colours $\Rightarrow$ Background Colour\ldots} option.
+\section{Importing Features from Databases}
+\label{featuresfromdb}
+Jalview supports feature retrieval from public databases either directly or {\sl
+via} the Distributed Annotation System (DAS\footnote{http://www.biodas.org/}).
+It includes built in parsers for Uniprot and ENA (or EMBL) records retrieved
+from the EBI. Sequences retrieved from these sources using the sequence fetcher (see
+Section \ref{fetchseq}) will already possess features.
-By default, the structure will be coloured in the same way as the associated
-sequence(s) in the alignment view from which it was launched. The structure can
-be coloured independently of the sequence by selecting an appropriate colour
-scheme from the {\sl Colours} menu. It can be coloured according to the
-alignment using the {\sl Colours $\Rightarrow$ By Sequence } option. The image
-in the structure viewer can be saved as an EPS or PNG with the {\sl File
-$\Rightarrow$ Save As $\Rightarrow$ \ldots} submenu, which also allows the raw
-data to be saved as PDB format. The mapping between the structure and the
-sequence (How well and which parts of the structure relate to the sequence) can
-be viewed with the {\sl File $\Rightarrow$ View Mapping} menu option.
+\subsection{Sequence Database Reference Retrieval}
+\label{fetchdbrefs}
+Jalview maintains a list of external database references for each sequence in
+an alignment. These are listed in a tooltip when the mouse is moved over the
+sequence ID when the {\sl View $\Rightarrow$ Sequence ID Tooltip $\Rightarrow$
+Show Database Refs } option is enabled. Sequences retrieved using the sequence
+fetcher will always have at least one database reference, but alignments
+imported from an alignment file generally have no database references.
-\subsubsection{Using the Jmol visualization interface }
+\subsubsection{Database References and Sequence Coordinate Systems}
-Jmol has a comprehensive set of selection and visualization functions that are
-accessed from the Jmol popup menu (by right-clicking in the Jmol window or by
-clicking the Jmol logo). Molecule colour and rendering style can be manipulated,
-and distance measurements and molecular surfaces can be added to the view. It
-also has its own ``Rasmol\footnote{See \url{http://www.rasmol.org}}-like'' scripting
-language, which is described elsewhere\footnote{Jmol Wiki:
-\url{http://wiki.jmol.org/index.php/Scripting}
+Jalview displays features in the local sequence's coordinate system which is
+given by its `start' and `end'. Any sequence features on the sequence will be
+rendered relative to the sequence's start position. If the start/end positions
+do not match the coordinate system from which the features were defined, then
+the features will be displayed incorrectly.
-Jmol Scripting reference:
-\url{http://www.stolaf.edu/academics/chemapps/jmol/docs/}}. Jalview utilises the
-scripting language to interact with Jmol and to store the state of a Jmol
-visualization within Jalview archives, in addition to the PDB data file
-originally loaded or retrieved by Jalview. To access the Jmol scripting
-environment directly, use the {\sl Jmol $\Rightarrow$ Console} menu option.
+\subsubsection{Viewing and Exporting a Sequence's Database Annotation}
-If you would prefer to use Jmol to manage structure colours, then select the
-{\sl Colours $\Rightarrow$ Colour with Jmol} option. This will disable any
-automatic application of colour schemes when new structure data is added, or
-when associated alignment views are modified.
+You can export all the database cross references and annotation terms shown in
+the sequence ID tooltip for a sequence by right-clicking and selecting the {\sl
+[Sequence ID] $\Rightarrow$ Sequence details \ldots} option from the popup menu.
+A similar option is provided in the {\sl Selection} sub-menu allowing you to
+obtain annotation for the sequences currently selected.
+\parbox[l]{3.4in}{
+The {\sl Sequence Details
+\ldots} option will open a window containing the same text as would be shown in
+the tooltip window, including any web links associated with the sequence. The
+text is HTML, and options on the window allow the raw code to be copied and
+pasted into a web page.}
+\parbox[c]{3in}{
+\centerline{\includegraphics[width=2.2in]{images/seqdetailsreport.pdf}}}
-\exercise{Viewing Structures}{\label{viewingstructex}
-\exstep{Load the alignment at
-\textsf{http://www.jalview.org/examples/exampleFile.jar}. Right-click on the
-sequence ID label for any of the sequences (e.g. {\sl FER1\_SPIOL}) to bring up
-the context menu. Select {\sl FER1\_SPIOL $\Rightarrow$ Structure $\Rightarrow$
-Associate Structure with Sequence $\Rightarrow$ Discover
-PDB IDs}. Jalview will now attempt to find PDB structures for the sequences in
-the alignment.
-% JBP Note: Bug JAL-1238 needs to be fixed ASAP
+\subsubsection{Automatically Discovering a Sequence's Database References}
+Jalview includes a function to automatically verify and update each sequence's
+start and end numbering against any of the sequence databases that the {\sl
+Sequence Fetcher} has access to. This function is accessed from the {\sl
+Webservice $\Rightarrow$ Fetch DB References} sub-menu in the Alignment
+window. This menu allows you to query either the set of {\sl Standard
+Databases}, which includes EMBL, Uniprot, the PDB, and the currently selected
+DAS sequence sources, or just a specific datasource from one of the submenus.
+When one of the entries from this menu is selected, Jalview will use the ID
+string from each sequence in the alignment or in the currently selected set to
+retrieve records from the external source. Any sequences that are retrieved are
+matched against the local sequence, and if the local sequence is found to be a
+sub-sequence of the retrieved sequence then the local sequence's start/end
+numbering is updated. A new database reference mapping is created, mapping the
+local sequence to the external database, and the local sequence inherits any
+additional annotation retrieved from the database sequence.
-{\bf Note:} If you are using Jalview v2.8 - use the {\sl Uniprot } source from the {\sl Web services $\Rightarrow$ Fetch DB References $\Rightarrow$ ..} submenu of the Alignment Window to retrieve the PDB IDs. }
-\exstep{ Right-click on the sequence ID for {\sl FER1\_SPIOL}.
-Select { \sl FER1\_SPIOL $\Rightarrow$ Structure $\Rightarrow$ View Structure
-$\Rightarrow$ 1A70}. A structure viewing window appears. Rotate the molecule by clicking and dragging in the structure viewing box. Zoom with the mouse scroll wheel. } \exstep{Roll the mouse cursor along the {\sl FER1\_SPIOL} sequence in the alignment. Note that if a residue in the sequence maps to one in the structure, a label will appear next to that residue in the structure viewer. Move the mouse over the structure. Placing the mouse over a part of the structure will bring up a tool tip indicating the name and number of that residue. The corresponding residue in the sequence is highlighted in black. Clicking the alpha carbon toggles the highlight and residue label on and off. Try this by clicking on a set of three or four adjacent residues so that the labels are persistent, then finding where they are in the sequence. }
-\exstep{Select {\sl Colours $\Rightarrow$ Background Colour\ldots} from the structure viewer menu and choose a suitable colour. Press OK to apply this. Select {\sl File $\Rightarrow$ Save As $\Rightarrow$ PNG} and save the image. View this with a suitable program. }
-\exstep{Select {\sl File $\Rightarrow$ View Mapping} from the structure viewer menu. A new window opens showing the residue by residue alignment between the sequence and the structure.}
-\exstep{Select {\sl File $\Rightarrow$ Save $\Rightarrow$ PDB file} and choose a new filename to save the PDB file. Once the file is saved, open the location in your file browser (or explorer window) and drag the PDB file that you just saved on to the Jalview desktop (or load it from the {\sl Jalview Desktop $\Rightarrow$ Input Alignment $\Rightarrow$ From File } menu). Verify that you can open and view the associated structure from the sequence ID pop-up menu's {\sl Structure } submenu in the new alignment window.}
+The database retrieval process terminates when a valid mapping is found for a
+sequence, or if all database queries failed to retrieve a matching sequence.
+Termination is indicated by the disappearance of the moving progress indicator
+on the alignment window. A dialog box may be shown once it completes which
+lists sequences for which records were found, but the sequence retrieved from
+the database did not exactly contain the sequence given in the alignment (the
+{\sl ``Sequence not 100\% match'' dialog box}).
+
+\exercise{Retrieving Database References}{
+\exstep{Load the example alignment at http://www.jalview.org/tutorial/alignment.fa}
+\exstep{Verify that there are no database references for the sequences by first
+checking that the {\sl View $\Rightarrow$ Sequence ID Tooltip $\Rightarrow$ Show
+Database Refs} option is selected, and then mousing over each sequence's ID.}
+\exstep{Use the {\sl Webservice $\Rightarrow$ Fetch DB References} menu option to retrieve database IDs for the sequences.}
+\exstep{Examine the tooltips for each sequence in the alignment as the retrieval progresses - note the appearance of new database references.}
+\exstep{Once the process has finished, save the alignment as a Jalview Project.}
+\exstep{Now close all the windows and open the project again, and verify that the database references and sequence features are still present on the alignment}
-\exstep{Right click on the structure to bring up the Jmol window. Explore the menu options. Try to change the style of molecular display - by first using the {\sl Jmol $\Rightarrow$ Select (n) $\Rightarrow$ All} command (where {\sl n} is the number of residues selected), and then the {\sl Jmol $\Rightarrow$ Style $\Rightarrow$ Scheme $\Rightarrow$ Ball and Stick} command.}
-\exstep{Use the {\sl File $\Rightarrow$ Save As .. } function to save the alignment as a Jalview Project. Now close the alignment and the structure view, and load the project file you just saved.
+\exstep{View the {\sl Sequence details \ldots} report for the FER1\_SPIOL sequence and for the whole alignment. Which sequences have web links associated with them ?}
-Verify that the Jmol display is as it was when you just saved the file.}
}
-\subsection{Superimposing structures}
-\label{superposestructs}
-Many comparative biomolecular analysis investigations aim to determine if the
-biochemical properties of a given molecule are significantly different to its
-homologues. When structure data is available, comparing the shapes of molecules
-by superimposing them enables substructure that may impart different behaviour
-to be quickly identified. The identification of optimal 3D superpositions
-involves aligning 3D data rather than sequence symbols, but the result can
-still be represented as a sequence alignment, where columns indicate positions
-in each molecule that should be superposed to recreate the optimal 3D alignment.
+\subsection{Retrieving Features {\sl via} DAS}
+\label{dasfretrieval}
+Jalview includes a client to retrieve features from DAS annotation servers. To
+retrieve features, select {\sl View $\Rightarrow$ Feature Settings\ldots} from the alignment window menu. Select the {\sl DAS Settings} tab in the Sequence Feature Settings Window (Figure \ref{das}). A list of DAS sources compiled from the currently configured DAS registry\footnote{By default, this will be the major public DAS server registry maintained by the Sanger Institute: http://www.dasregistry.org} is shown in the left hand pane. Highlighting an entry on the left brings up information about that source in the right hand panel.
-Jalview can employ Jmol's 3D fitting routines\footnote{See
-\href{http://chemapps.stolaf.edu/jmol/docs/?ver=12.2$\#$compare}{http://chemapps.stolaf.edu/jmol/docs/?ver=12.2$\#$compare}
-for more information.} to recreate 3D structure superpositions based on the
-correspondences defined by one or more sequence alignments involving structures shown in the Jmol display. Superposition based on the currently displayed alignment view happens automatically if a
-structure is added to an existing Jmol display using the {\sl Structure
-$\Rightarrow$ View PDB Structure $\Rightarrow$ ..}. A new Jmol view containing
-superposed structures can also be created using the {\sl Structure
-$\Rightarrow$ View all {\bf N} PDB Structures} option (when {\bf {\sl N}}
-$>$ 1) if the current selection contains two or more sequences with associated
-structures.
+\begin{figure}[htbp]
+\begin{center}
+\includegraphics[width=2.5in]{images/das1.pdf}
+\includegraphics[width=2.5in]{images/das2.pdf}
+\caption{{\bf Retrieving DAS annotations.} DAS features are retrieved using the {\sl DAS Settings} tab (left) and their display customised using the {\sl Feature Settings} tab (right).}
+\label{das}
+\end{center}
+\end{figure}
-\subsubsection{Obtaining the RMSD for a superposition}
-The RMSD (Root Mean Square Deviation) is a measure of how similar the structures
-are when they are superimposed. Figure \ref{mstrucsuperposition} shows a
-superposition created during the course of Exercise \ref{superpositionex}. The
-parts of each molecule used to construct the superposition are rendered using
-the cartoon style, with other parts of the molecule drawn in wireframe. The Jmol
-console, which has been opened after the superposition was performed, shows the
-RMSD report for the superposition.
-Full information about the superposition is also output to the Jalview
-console.\footnote{The Jalview Java Console is opened from {\sl Tools
-$\Rightarrow$ Java Console} option in the Desktop's menu bar} This output also
-includes the precise atom pairs used to superpose structures.
+Select appropriate DAS sources as required then click on {\sl Fetch DAS
+Features}. If you know of additional sources not listed in the configured
+registry, then you may add them with the {\sl Add Local Source} button. Use
+the {\sl Authority},{\sl Type}, and {\sl Label} filters to restrict the list
+of sources to just those that will return features for the sequences in the
+alignment.
-\subsubsection{Choosing which part of the alignment is used for structural
-superposition} Jalview uses the visible part of each alignment view to define
-which parts of each molecule are to be superimposed. Hiding a column in a view
-used for superposition will remove that correspondence from the set, and will
-exclude it from the superposition and RMSD calculation.
-This allows the selection of specific parts of the alignment to be used for
-superposition. Only columns that define a complete set of correspondences for
-all structures will be used for structural superposition, and as a consequence,
-the RMSD values generated for each pair of structures superimposed can be
-directly compared.
+Following DAS feature retrieval, the {\sl Feature Settings} panel takes on a
+slightly different appearance (Figure \ref{das} (right)). Each data source is
+listed and groups of features from one data source can be selected/deselected
+by checking the labelled box at the top of the panel.
-In order to recompute a superposition after changing a view or editing the
-alignment, select the {\sl Jmol $\Rightarrow$ Align sequences } menu option. The {\sl
-Jmol $\Rightarrow$ Superpose with ..} submenu allows you to choose which of the
-associated alignments and views are to be used to create the set of
-correspondences. This menu is useful when composing complex superpositions
-involving multi-domain and multi-chain complexes, when correspondences may be
-defined by more than one alignment.
+\exercise{Retrieving Features with DAS}{
+\label{dasfeatretrexcercise}
+\exstep{Load the alignment at
+\textsf{http://www.jalview.org/tutorial/alignment.fa}. Select {\sl View
+$\Rightarrow$ Feature Settings \ldots} from the alignment window menu. Select
+the {\sl DAS Settings} tab. A long list of available DAS sources is listed.
+Select a small number, eg Uniprot, DSSP, signalP and netnglyc. Click.
+A window may prompt whether you wish Jalview to fetch DAS features. Click {\sl
+Yes}.
+Jalview will start retrieving features. As features become available they will be mapped onto the alignment. }
+\exstep{If Jalview is taking too long to retrieve features, the process can be cancelled with the {\sl Cancel Fetch} button.
+Rolling the mouse cursor over the sequences reveals a large number of features annotated in the tool tip.
+Close the Sequence Feature Settings window. }
+\exstep{Move the mouse over the sequence ID panel.
+Non-positional features such as literature references and protein localisation predictions are given in the tooltip, below any database cross references associated with the sequence.}
+\exstep{Search through the alignment to find a feature with a link symbol next to it.
+Right click to bring up the alignment view popup menu, and find a corresponding entry in the {\sl Link } sub menu. }
+% TODO this doesn't work ! \includegraphics[width=.3in]{images/link.pdf}
-Note that these menu options appear when you have two or more structures in one Jmol viewer.
+\exstep{
+Select {\sl View $\Rightarrow$ Feature Settings\ldots} to reopen the Feature Settings window. All the loaded feature types should now be displayed. Those at the top of the list are drawn on top of those below, obscuring them in the alignment view where they overlap. Move the feature settings window so that the alignment is visible and uncheck some of the feature types by clicking the tick box in the display column. Observe how the alignment display changes. Note that unselected feature types do not appear in the tool tip.
+}
+\exstep{Reorder the features by dragging feature types up and down the order in the Feature Settings panel. e.g. Click on {\sl CHAIN} then move the mouse downwards to drag it below {\sl DOMAIN}. Note that {\sl DOMAIN} is now shown on top of {\sl CHAIN} in the alignment window. Drag {\sl METAL} to the top of the list. Observe how the cysteine residues are now highlighted as they have a {\sl METAL} feature associated with them.
+}
+\exstep{Press the {\sl Optimise Order} button. The features will be ordered according to increasing length, placing features that annotate shorter regions of sequence higher on the display stack.}
+\exstep{Select {\sl File $\Rightarrow$ Export Features\ldots} from the Alignment window. You can choose to export the retrieved features as a GFF file, or Jalview's own Features format.
+% TODO: describe working with features files and GFF
+}
+}
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=5.5in]{images/fdxsuperposition.pdf}
-\caption{{\bf Superposition of two ferredoxin structures.} The alignment on the
-left was used by jalview to superpose structures associated with the
-FER1\_SPIOL and FER1\_MAIZE sequences in the alignment. Parts of each structure
-used for superposition are rendered as a cartoon, the remainder rendered in
-wireframe. The RMSD between corresponding positions in the structures before and
-after the superposition is shown in the Jmol console.}
-\label{mstrucsuperposition}
-\end{center}
-\end{figure}
+\subsubsection{The Fetch Uniprot IDs Dialog Box}
+\label{discoveruniprotids}
+If any sources are selected which refer to Uniprot coordinates as their reference system,
+then you may be asked if you wish to retrieve Uniprot IDs for your sequence. Pressing OK instructs Jalview to verify the sequences against Uniprot records retrieved using the sequence's ID string. This operates in much the same way as the {\sl Web Service $\Rightarrow$ Fetch Database References } function described in Section \ref{fetchdbrefs}. If a sequence is verified, then the start/end numbering will be adjusted to match the Uniprot record to ensure that features retrieved from the DAS source are rendered at the correct position.
-\exercise{Aligning structures using the ferredoxin
-sequence alignment.}{\label{superpositionex}
+\subsubsection{Rate of Feature Retrieval}
+Feature retrieval can take some time if a large number of sources is selected and if the alignment
+contains a large number of sequences. This is because Jalview only queries a particular DAS source with one sequence at a time, to avoid overloading it. As features are retrieved, they are immediately added to the current alignment view. The retrieved features are shown on the sequence and can be customised as described previously.
-\exstep{Continue with the Jalview project created in exercise
-\ref{viewingstructex}. Use the {\sl Discover PDB IDs} function to retrieve PDB
-IDs associated with the FER1\_MAIZE sequence.}
-\exstep{Once discovery has completed, use the {\sl
-View PDB Structure} submenu to view the PDB file associated with FER1\_MAIZE.
-Jalview will give you the option of aligning the structure to the one already
-open. To superimpose the structure associated with FER1\_MAIZE with the one
-associated with FER1\_SPIOL, press the {\bf Yes} button.
-{\sl The Jmol view will update to show both structures, and one will be
-moved on to the other. If this doesn't happen, use the Align function in the Jmol submenu}}
-\exstep{Create a new view on the alignment, and hide all but columns 121
-through to 132.}
-\exstep{Use the {\sl Jmol} submenu to
-recompute the superposition using just columns 121-132 of the alignment.
+\subsection{Colouring Features by Score or Description
+Text}
+\label{featureschemes}
+Sometimes, you may need to visualize the differences in information carried by
+sequence features of the same type. This is most often the case when features
+of a particular type are the result of a specific type of database query or calculation. Here, they may also carry information within their textual description, or most commonly for calculations, a score related to the property being investigated. Jalview can shade sequence
+features using a graduated colourscheme in order to highlight these variations.
+In order to apply a graduated scheme to a feature type, select the `Graduated
+colour' entry in the Sequence {\sl Feature Type}'s popup menu, which is opened by
+right-clicking the {\sl Feature Type} or {\sl Color} in the {\sl Sequence Feature Settings} dialog box. Two types
+of colouring styles are currently supported: the default is quantitative
+colouring, which shades each feature based on its score, with the highest
+scores receiving the `Max' colour, and the lowest scoring features coloured
+with the `Min' colour. Alternately, you can select the `Colour by label'
+option to create feature colours according to the description text associated
+with each feature. This is useful for general feature types - such as
+Uniprot's `DOMAIN' feature - where the actual type of domain is given in the
+feature's description.
-{\sl Note how the molecules shift position when superposed using a short part of
-the two structures.}}
-\exstep{Compare the initial and final RMSDs for superimposing molecules with
-the small section and with the whole alignment. Which view do you think give the
-best 3D superposition, and why ?} }
+Graduated feature colourschemes can also be used to exclude low or
+high-scoring features from the alignment display. This is done by choosing your
+desired threshold type (either above or below), using the drop-down menu in the
+dialog box. Then, adjust the slider or enter a value in the text box to set the
+threshold for displaying this type of feature.
-\subsection{Colouring structure data associated with multiple alignments and views}
-Normally, the original view from which a particular structure view was
-opened will be the one used to colour structure data. If alignments involving
-sequences associated with structure data shown in a Jmol have multiple views, Jalview gives you full control
-over which alignment, or alignment view, is used to colour the structure
-display. Sequence-structure colouring associations are
-changed {\sl via} the {\sl View $\Rightarrow$ Colour by ..} menu, which lists all
-views associated with data shown in the embedded Jmol view. A tick is shown beside
-views currently used as colouring source, and moving the
-mouse over each view will bring it to the front of the alignment display,
-allowing you to browse available colour sources prior to selecting one. If the
-{\sl Select many views} option is selected, then multiple views can be selected as sources for colouring the
-structure data. {\sl Invert selection} and {\sl Select all views} options are also provided to quickly change between multi-view selections.
+The feature settings dialog box allows you to toggle between a graduated and
+simple feature colourscheme using the pop-up menu for the feature type. When a
+graduated scheme is applied, it will be indicated in the colour column for
+that feature type - with coloured blocks or text to indicate the colouring
+style and a greater than ($>$) or less than ($<$) symbol to indicate when a
+threshold has been defined.
-Note that the {\sl Select many views} option is useful if you have different
-views that colour different areas or domains of the alignment. This option is
-further explored in Section \ref{complexstructurecolours}.
+\subsection{Using Features to Re-order the Alignment}
+\label{featureordering}
+The presence of sequence features on certain sequences or in a particular
+region of an alignment can quantitatively identify important trends in
+the aligned sequences. In this case, it is more useful to
+re-order the alignment based on the number of features or their associated scores, rather than simply re-colour the aligned sequences. The sequence feature settings
+dialog box provides two buttons: `Seq sort by Density' and `Seq sort by
+Score', that allow you to reorder the alignment according to the number of
+sequence features present on each sequence, and also according to any scores
+associated with a feature. Each of these buttons uses the currently displayed
+features to determine the ordering, but
+if you wish to re-order the alignment using a single type of feature, then you can do this from the {\sl Feature Type}'s
+popup menu. Simply right-click the type's style in the Sequence Feature Settings dialog
+box, and select one of the {\sl Sort by Score} and {\sl Sort by Density}
+options to re-order the alignment. Finally, if a specific region is selected,
+then only features found in that region of the alignment will be used to
+create the new alignment ordering.
+% \exercise{Shading and Sorting Alignments using Sequence Features}{
+% \label{shadingorderingfeatsex}
+%
+% This exercise is currently not included in the tutorial because no DAS servers
+% currently exist that yield per-residue features for any Uniprot sequence.
+%
+% \exstep{Re-load the alignment from \ref{dasfeatretrexcercise}.
+% }
+% \exstep{Open the
+% feature settings panel, and, after first clearing the current
+% selection, press the {\em Seq Sort by Density} button a few times.}
+% \exstep{Use the DAS fetcher to retrieve the Kyte and Doolittle Hydrophobicity
+% scores for the protein sequences in the alignment.
+% {\sl Hint: the nickname for the das source is `KD$\_$hydrophobicity'.}}
+% \exstep{Change the feature settings so only the hydrophobicity features are
+% displayed. Mouse over the annotation and also export and examine the GFF and
+% Jalview features file to better understand how the hydrophobicity measurements
+% are recorded.}
+% \exstep{Apply a {\sl Graduated Colour} to the hydrophobicity annotation to
+% reveal the variation in average hydrophobicity across the alignment.}
+% \exstep{Select a range of alignment columns, and use one of the sort by feature buttons to order the alignment according to that region's average
+% hydrophobicity.}
+% \exstep{Save the alignment as a project, for use in exercise
+% \ref{threshgradfeaturesex}.} }
+%
+% \exercise{Shading alignments with combinations of graduated feature
+% colourschemes}{
+% \label{threshgradfeaturesex}
+% \exstep{Reusing the annotated alignment from exercise
+% \ref{shadingorderingfeatsex}, experiment with the colourscheme threshold to
+% highlight the most, or least hydrophobic regions. Note how the {\sl Colour} icon for the {\sl Feature Type} changes when you change the threshold type and press OK.}
+% \exstep{Change the colourscheme so
+% that features at the threshold are always coloured grey, and the most
+% hydrophobic residues are coloured red, regardless of the threshold value
+% ({\em hint - there is a switch on the dialog to do this for you}).}
+% \exstep{Enable the Uniprot {\em chain} annotation in the feature settings
+% display and re-order the features so it is visible under the hydrophobicity
+% annotation.}
+% \exstep{Apply a {\sl Graduated Colour} to the {\em chain}
+% annotation so that it distinguishes the different canonical names associated
+% with the mature polypeptide chains.}
+% \exstep{Export the alignment's sequence features using the Jalview and GFF file formats, to see how the different types of graduated feature
+% colour styles are encoded. }
+% }
+
+\subsection{Creating Sequence Features}
+Sequence features can be created simply by selecting the area in a sequence (or sequences) to form the feature and selecting {\sl Selection $\Rightarrow$ Create Sequence Feature } from the right-click context menu (Figure \ref{features}). A dialogue box allows the user to customise the feature with respect to name, group, and colour. The feature is then associated with the sequence. Moving the mouse over a residue associated with a feature brings up a tool tip listing all features associated with the residue.
\begin{figure}[htbp]
\begin{center}
-\includegraphics[width=5.5in]{images/mviewstructurecol.pdf}
-\caption{{\bf Choosing a different view for colouring a structure display}
-Browsing the {\sl View $\Rightarrow$ Colour by ..} menu provides full control
-of which alignment view is used to colour structures when the {\sl Colours
-$\Rightarrow$ By Sequence} option is selected.}
-\label{mviewstructurecol}
+\includegraphics[width=2in]{images/feature1.pdf}
+\includegraphics[width=2.5in]{images/feature2.pdf}
+\includegraphics[width=1.5in]{images/feature3.pdf}
+\caption{{\bf Creating sequence features.} Features can readily be created from selections via the context menu and are then displayed on the sequence. }
+\label{features}
\end{center}
\end{figure}
-\subsubsection{Colouring complexes}
-\label{complexstructurecolours}
-The ability to control which multiple alignment view is used to colour
-structural data is essential when working with data relating to
-multidomain biomolecules and complexes.
+Creation of features from a selection spanning multiple sequences results in the creation of one feature per sequence.
+Each feature remains associated with its own sequence.
-In these situations, each chain identified in the structure may have a different
-evolutionary history, and a complete picture of functional variation can
-only be gained by integrating data from different alignments on the same
-structure view. An example of this is shown in Figure
-\ref{mviewalcomplex}, based on data from Song et. al\footnote{Structure of
-DNMT1-DNA Complex Reveals a Role for Autoinhibition in Maintenance DNA Methylation. Jikui Song, Olga Rechkoblit, Timothy H. Bestor, and Dinshaw J. Patel.
-{\sl Science} 2011 {\bf 331} 1036-1040
-\href{http://www.sciencemag.org/content/331/6020/1036}{DOI:10.1126/science.1195380}}
+\subsection{Customising Feature Display}
+
+Feature display can be toggled on or off by selecting the {\sl View
+$\Rightarrow$ Show Sequence Features} menu option. When multiple features are
+present it is usually necessary to customise the display. Jalview allows the
+display, colour, rendering order and transparency of features to be modified
+{\sl via} the {\sl View $\Rightarrow$ Feature Settings\ldots} menu option. This
+brings up a dialogue window (Figure \ref{custfeat}) which allows the
+visibility of individual feature types to be selected, colours changed (by
+clicking on the colour of each sequence feature type) and the rendering order
+modified by dragging feature types to a new position in the list. Dragging the
+slider alters the transparency of the feature rendering. The Feature
+Settings dialog also includes functions for more advanced feature shading
+schemes and buttons for sorting the alignment according to the distribution of
+features. These capabilities are described further in sections
+\ref{featureschemes} and \ref{featureordering}.
\begin{figure}[htbp]
\begin{center}
-\includegraphics[]{images/mchainstructureview.pdf}
-\caption{{\bf The biological assembly of Mouse DNA Methyltransferase-1 coloured
-by Pfam alignments for its major domains} Alignments for each domain within the
-Uniprot sequence DNMT1\_MOUSE have been used to visualise sequence conservation
-in each component of this protein-DNA complex. Instructions for recreating this figure are given in exercise \ref{dnmtcomplexex}. }
-\label{mviewalcomplex}
+\includegraphics[width=4in]{images/features4.pdf}
+\caption{{\bf Multiple sequence features.} An alignment with JPred secondary structure prediction annotation below it, and many sequence features overlaid onto the aligned sequences. The tooltip lists the features annotating the residue below the mouse-pointer.}
\end{center}
\end{figure}
-\exercise{Colouring a protein complex to explore domain-domain interfaces}{\label{dnmtcomplexex}
-
-\exstep{Download the PDB file at
-\textsf{\url{http://www.jalview.org/tutorial/DNMT1\_MOUSE.pdb}} to your desktop. This
-is the biological unit for PDB ID 3pt6, as identified by the PDBe's PISA server.}
-
-\exstep{Launch the Jalview desktop and ensure you have at least 256MB of
-free memory available.
-
-{\sl Use the following webstart link:
-\href{http://www.jalview.org/webstart/jalview_1G.jnlp}{http://www.jalview.org/webstart/jalview\_1G.jnlp}}.}
-\exstep{Retrieve the following
-{\bf full} PFAM alignments: PF02008, PF00145, PF01426 (make sure you select the {\sl PFAM {\bf (Full)}} source). These will each be retrieved into their own alignment window.}
-\exstep{Drag the structure you downloaded in
-step 1 onto one of the alignments to associate it with the mouse sequence in
-that Pfam domain family.}
-\exstep{For every DNMT1\_MOUSE sequence in the alignment, use the sequence
-ID popup menu's {\sl Structure} submenu to view the DNMT1\_MOUSE structure for the associated mouse sequence. When given the option, {\bf view all of the structures in the same Jmol viewer}. Check the contents of the {\sl View $\Rightarrow$ Colour by ..} submenu to see what alignments can be used to
-colour the sequence.}
-\exstep{Repeat the previous two steps for each of
-the other alignments. In each case, when performing the `View DNMT1\_MOUSE.pdb'
-step, Jalview will ask if you wish to create a new Jmol view. You should
-respond `No', {\bf ensuring that each sequence fragment is associated with the same Jmol view}.}
-\exstep{Pick a different colourscheme for each alignment, and use the {\sl
-Colour by ..} submenu to ensure they are all used to colour the complex shown
-in the Jmol window.}
-\exstep{The final step needed to reproduce the shading in Figure
-\ref{mviewalcomplex} is to use the {\sl Colour $\Rightarrow$ By
-Annotation } option in each alignment window to shade the alignment by the
-{\bf Conservation} annotation row. This function was described in section
-\ref{colourbyannotation}.
+\begin{figure}[htbp]
+\begin{center}
+\includegraphics[width=4in]{images/features5.pdf}
+\caption{{\bf Customising sequence features.} Features can be recoloured, switched on or off and have the rendering order changed. }
+\label{custfeat}
+\end{center}
+\end{figure}
-Ensure that you first disable the {\sl View $\Rightarrow$ Show Features} menu option, or you may not see any colour changes in the associated structure.
+\subsection{Sequence Feature File Formats}
-{\sl Note: Choose a different shading scheme for each
-alignment so that the regions of strong physicochemical conservation are highlighted. This
-kind of shading will reveal conserved regions of interaction between domains
-in the structure.}}
-\exstep{Save your work as a Jalview project and verify that it can be opened again by starting another Jalview Desktop instance, and dragging the saved project into the desktop window.}
+Jalview supports the widely used GFF tab delimited format\footnote{see
+http://www.sanger.ac.uk/resources/software/gff/spec.html} and its own Jalview
+Features file format for the import of sequence annotation. Features and
+alignment annotation are also extracted from other formats such as Stockholm,
+and AMSA. URL links may also be attached to features. See the online
+documentation for more details of the additional capabilities of the Jalview
+features file.
-{\sl Note: This exercise relies on new features introduced in Jalview 2.7. If
-you notice any strange behaviour when trying out this exercise, it may be a
-bug (see
-\href{http://issues.jalview.org/browse/JAL-1008}{http://issues.jalview.org/browse/JAL-1008}
-for one relating to highlighting of positions in the alignment window).} }
+\exercise{Creating Features}{
+\exstep{Open the alignment at \textsf{http://www.jalview.org/tutorial/alignment.fa}.
+We know that the Cysteine residues at columns 97, 102, 105 and 135 are involved in
+iron binding so we will create them as features. Navigate to column 97, sequence 1.
+Select the entire column by clicking in the ruler bar. Then right-click on the selection
+to bring up the context menu and select {\sl Selection $\Rightarrow$ Create Sequence Feature}.
+A dialogue box will appear.
+}
+\exstep{
+Enter a suitable Sequence Feature Name (e.g. ``Iron binding site") in the
+appropriate box. Click on the Feature Colour bar to change the colour if
+desired, add a short description (``One of four Iron binding Cysteines") and press OK. The features will then appear on the sequences. } \exstep{Roll the mouse cursor over the new features. Note that the position given in the tool tip is the residue number, not the column number. To demonstrate that there is one feature per sequence, clear all selections by pressing [ESC] then insert a gap in sequence 3 at column 95. Roll the mouse over the features and you will see that the feature has moved with the sequence. Delete the gap you created.
+}
+\exstep{
+Add a similar feature to column 102. When the feature dialogue box appears, clicking the Sequence Feature
+Name box brings up a list of previously described features. Using the same Sequence Feature Name allows the features to be grouped.}
+\exstep{Select {\sl View $\Rightarrow$ Feature Settings\ldots} from the
+alignment window menu. The Sequence Feature Settings window will appear. Move
+this so that you can see the features you have just created. Click the check
+box for ``Iron binding site" under {\sl Display} and note that display of this
+feature type is now turned off. Click it again and note that the features are
+now displayed. Close the sequence feature settings box by clicking OK or
+Cancel.} }
-\section{Analysis of alignments}
-\label{alignanalysis}
-Jalview provides support for sequence analysis in two ways. A number of
-analytical methods are `built-in', these are accessed from the {\sl Calculate}
-alignment window menu. Computationally intensive analyses are run outside
-Jalview {\sl via} web services - these are typically accessed {\sl via} the {\sl
-Web Service} menu, and described in \ref{jvwebservices} and subsequent sections.
-In this section, we describe the built-in analysis capabilities common to both
-the Jalview Desktop and the JalviewLite applet.
-
-\subsection{PCA}
-This calculation creates a spatial representation of the similarities within the
-current selection or the whole alignment if no selection has been made. After
-the calculation finishes, a 3D viewer displays the each sequence as a point in
-3D `similarity space'. Sets of similar sequences tend to lie near each other in
-this space.
+\chapter{Multiple Sequence Alignment}
+\label{msaservices}
+Sequences can be aligned using a range of algorithms provided by JABA web
+services. These include ClustalW\footnote{{\sl ``CLUSTAL W: improving the
+sensitivity of progressive multiple sequence alignment through sequence
+weighting, position specific gap penalties and weight matrix choice."} Thompson
+JD, Higgins DG, Gibson TJ (1994) {\sl Nucleic Acids Research} {\bf 22},
+4673-80}, Muscle\footnote{{\sl ``MUSCLE: a multiple sequence alignment method
+with reduced time and space complexity"} Edgar, R.C.
+(2004) {\sl BMC Bioinformatics} {\bf 5}, 113}, MAFFT\footnote{{\sl ``MAFFT: a
+novel method for rapid multiple sequence alignment based on fast Fourier
+transform"} Katoh, K., Misawa, K., Kuma, K. and Miyata, T. (2002) {\sl Nucleic
+Acids Research} {\bf 30}, 3059-3066. and {\sl ``MAFFT version 5:
+improvement in accuracy of multiple sequence alignment"} Katoh, K., Kuma, K.,
+Toh, H. and Miyata, T. (2005) {\sl Nucleic Acids Research} {\bf 33}, 511-518.},
+ProbCons,\footnote{PROBCONS: Probabilistic Consistency-based Multiple Sequence
+Alignment.
+Do, C.B., Mahabhashyam, M.S.P., Brudno, M., and Batzoglou, S.
+(2005) {\sl Genome Research} {\bf 15} 330-340.} T-COFFEE\footnote{T-Coffee:
+A novel method for multiple sequence alignments. (2000) Notredame, Higgins and
+Heringa {\sl JMB} {\bf 302} 205-217} and Clustal Omega.\footnote{Fast, scalable
+generation of high-quality protein multiple sequence alignments using Clustal
+Omega. Sievers F, Wilm A, Dineen DG, Gibson TJ, Karplus K, Li W, Lopez R,
+McWilliam H, Remmert M, Soding J, Thompson JD, Higgins DG (2011) {\sl Molecular
+Systems Biology} {\bf 7} 539
+\href{http://dx.doi.org/10.1038/msb.2011.75}{doi:10.1038/msb.2011.75}} Of these,
+T-COFFEE is the slowest, but also the most accurate. ClustalW is historically
+the most widely used. Muscle is faster than ClustalW and probably the most
+accurate for smaller alignments and MAFFT is probably the best for large
+alignments, however {\bf Clustal Omega}, which was released in 2011, is
+arguably the fastest and most accurate tool for protein multiple alignment.
+
+
+To run an alignment web service, select the appropriate method from the {\sl
+Web Service $\Rightarrow$ Alignment $\Rightarrow$ \ldots} submenu (Figure
+\ref{webservices}). For each service you may either perform an alignment with
+default settings, use one of the available presets, or customise the parameters
+with the `{\sl Edit and Run ..}' dialog box. Once the job is submitted, a
+progress window will appear giving information about the job and any errors that
+occur. After successful completion of the job, a new alignment window is opened
+with the results, in this case an alignment. By default, the new alignment will be
+ordered in the same way as the input sequences. Note: many alignment
+programs re-order the input during their analysis and place homologous
+sequences close together, the MSA algorithm ordering can be recovered
+using the `Algorithm ordering' entry within the {\sl Calculate $\Rightarrow$
+Sort } sub menu.
+
+\subsubsection{Realignment}
+The re-alignment option is currently only supported by ClustalW and Clustal
+Omega. When performing a re-alignment, Jalview submits the current selection to
+the alignment service complete with any existing gaps. This approach is useful
+when one wishes to align additional sequences to an existing alignment without
+any further optimisation to the existing alignment. The re-alignment service
+provided by ClustalW in this case is effectively a simple form of profile
+alignment.
+
+
+\begin{figure}[htbp]
+\begin{center}
+\parbox[c]{1.5in}{\includegraphics[width=1.5in]{images/ws1.pdf}}
+\parbox[c]{2.5in}{\includegraphics[width=2.5in]{images/ws2.pdf}}
+\parbox[c]{2in}{\includegraphics[width=2in]{images/ws3.pdf}}
+\caption{{\bf Multiple alignment via web services} The appropriate method is
+selected from the menu (left), a status box appears (centre), and the results
+appear in a new window (right).}
+\label{webservices}
+\end{center}
+\end{figure}
+
+
+
+
+\exercise{Multiple Sequence Alignment}{
+\exstep{ Close all windows and open the alignment at {\sf
+http://www.jalview.org/tutorial/unaligned.fa}. Select {\sl
+Web Service $\Rightarrow$ Alignment $\Rightarrow$ Muscle with Defaults}.
+A window will open giving the job status. After a short time, a second window will open
+ with the results of the alignment.}
+ \exstep{Return to the first sequence alignment window by clicking on
+ the window, and repeat using Clustal and MAFFT (from the {\sl Web
+ Service $\Rightarrow$ Alignment} menu) on the same initial alignment. Compare them and
+ you should notice small differences. }
+\exstep{Select the last three sequences in the MAFFT alignment, and de-align them
+with {\sl Edit $\Rightarrow$ Remove All Gaps}. Press [ESC] to deselect them and then
+submit the view for re-alignment with Clustal.}
+\exstep{Use [CTRL]-Z to recover the alignment of the last three sequences in the MAFFT alignment.
+Once the Clustal re-alignment has completed, compare the results of re-alignment of the
+three sequences with their alignment in the original MAFFT result.}
+\exstep{Select columns 60 to 125 in the original MAFFT alignment and hide them.
+Select {\sl Web Services $\Rightarrow$ Alignment $\Rightarrow$ Mafft with Defaults} to
+submit the visible portion of the alignment to MAFFT. When the web service job pane appears,
+note that there are now two alignment job status panes shown in the window.}
+\exstep{When the MAFFT job has finished, compare the alignment of the N-terminal visible
+region in the result with the corresponding region of the original alignment. If you wish,
+select and hide a few more columns in the N-terminal region, and submit the alignment to the
+service again and explore the effect of local alignment on the non-homologous parts of the
+N-terminal region.}
+}
+
+\subsubsection{Alignments of Sequences that include Hidden Regions}
+
+If the view or selected region that is submitted for alignment contains hidden
+regions, then {\bf only the visible sequences will be submitted to the service}.
+Furthermore, each contiguous segment of sequences will be aligned independently
+(resulting in a number of alignment `subjobs' appearing in the status window).
+Finally, the results of each subjob will be concatenated with the hidden regions
+in the input data prior to their display in a new window. This approach ensures
+that 1) hidden column boundaries in the input data are preserved in the
+resulting alignment - in a similar fashion to the constraint that hidden columns
+place on alignment editing (see Section \ref{lockededits} and 2) hidden
+columns can be used to preserve existing parts of an alignment whilst the
+visible parts are locally refined.
+
+
+\subsection{Customising the Parameters used for Alignment}
+
+JABA web services allow you to vary the parameters used when performing a
+bioinformatics analysis. For JABA alignment services, this means you are
+usually able to modify the following types of parameters:
+\begin{list}{$\bullet$}{}
+\item Amino acid or nucleotide substitution score matrix
+\item Gap opening and widening penalties
+\item Types of distance metric used to construct guide trees
+\item Number of rounds of re-alignment or alignment optimisation
+\end{list}
+
+
+\subsubsection{Getting Help on the Parameters for a Service}
+Each parameter available for a method usually has a short description, which
+Jalview will display as a tooltip, or as a text pane that can be opened under
+the parameter's controls. In the parameter shown in Figure
+\ref{clustalwparamdetail}, the description was opened by selecting the button on the left hand side. Online help for the
+service can also be accessed, by right clicking the button and selecting a URL
+from the pop-up menu that will open.
+
+\begin{figure}[htbp]
+\begin{center}
+\includegraphics[width=2.5in]{images/clustalwparamdetail.pdf}
+\caption{{\bf ClustalW parameter slider detail}. From the ClustalW {\sl Clustal $\Rightarrow$ Edit settings and run ...} dialog box. }
+\label{clustalwparamdetail}
+\end{center}
+\end{figure}
+
+\subsection{Alignment Presets}
+The different multiple alignment algorithms available from JABA vary greatly in
+the number of adjustable parameters, and it is often difficult to identify what
+are the best values for the sequences that you are trying to align. For these
+reasons, each JABA service may provide one or more presets -- which are
+pre-defined sets of parameters suited for particular types of alignment
+problem. For instance, the Muscle service provides the following presets:
+\begin{list}{$\bullet$}{}
+\item Large alignments (balanced)
+\item Protein alignments (fastest speed)
+\item Nucleotide alignments (fastest speed)
+\end{list}
+
+The presets are displayed in the JABA web services submenu, and can also be
+accessed from the parameter editing dialog box, which is opened by selecting
+the `{\sl Edit settings and run ...}' option from the web services menu. If you have used
+a preset, then it will be mentioned at the beginning of the job status file shown
+in the web service job progress window.
+
+\subsubsection{Alignment Service Limits}
+Multiple alignment is a computationally intensive calculation. Some JABA server
+services and service presets only allow a certain number of sequences to be
+aligned. The precise number will depend on the server that you are using to
+perform the alignment. Should you try to submit more sequences than a service
+can handle, then an error message will be shown informing you of the maximum
+number allowed by the server.
+
+\subsection{User Defined Presets}
+Jalview allows you to create your own presets for a particular service. To do
+this, select the `{\sl Edit settings and run ...}' option for your service,
+which will open a parameter editing dialog box like the one shown in Figure
+\ref{jwsparamsdialog}.
+
+The top row of this dialog allows you to browse the existing presets, and
+when editing a parameter set, allows you to change its nickname. As you
+adjust settings, buttons will appear at the top of the parameters dialog that
+allow you to Revert or Update the currently selected user preset with your changes, Delete the current preset, or Create a new preset, if none exists with the given name. In addition to the parameter set name, you can also provide a short
+description for the parameter set, which will be shown in the tooltip for the
+parameter set's entry in the web services menu.
+
+\begin{figure}[htbc]
+\center{
+\includegraphics[width=3in]{images/jvaliwsparamsbox.pdf}
+\caption{{\bf Jalview's JABA alignment service parameter editing dialog box}.}
+\label{jwsparamsdialog} }
+\end{figure}
+
+\subsubsection{Saving Parameter Sets}
+When creating a custom parameter set, you will be asked for a file name to save
+it. The location of the file is recorded in the Jalview user preferences in the
+same way as a custom alignment colourscheme, so when Jalview is launched again,
+it will show your custom preset amongst the options available for running the
+JABA service.
+
+%
+% \exercise{Creating and using user defined presets}{\label{createandusepreseex}
+% \exstep{Import the file at
+% \textsf{http://www.jalview.org/tutorial/fdx\_unaligned.fa} into jalview.}
+% \exstep{Use the `{\slDiscover Database Ids}' function to recover the PDB cross
+% references for the sequences.}
+% \exstep{Align the sequences using the default ClustalW parameters.}
+% \exstep{Use the `{\sl Edit and run..}'
+% option to open the ClustalW parameters dialog box, and create a new preset using
+% the following settings:
+% \begin{list}{$\bullet$}{}
+% \item BLOSUM matrix (unchanged)
+% \item Gap Opening and End Gap penalties = 0.05
+% \item Gap Extension and Separation penalties = 0.05
+% \end{list}
+%
+% As you edit the parameters, buttons will appear on the dialog box
+% allowing you revert your changes or save your settings as a new parameter
+% set.
+%
+% Before you save your settings, remeber to give them a meaningful name by editing
+% the text box at the top of the dialog box.
+% }
+% \exstep{Repeat the alignment using your new parameter set by selecting it from
+% the {\sl ClustalW Presets menu}.}
+% \exstep{These sequences have PDB structures associated with them, so it is
+% possible to compare the quality of the alignments.
+%
+% Use the {\sl View all {\bf N}
+% structures} option to calculate the superposition of 1fdn on 1fxd for both
+% alignments (refer to section \ref{superposestructs} for instructions). Which
+% alignment gives the best RMSD ? }
+% \exstep{Apply the same alignment parameter settings to the example alignment
+% (available from \textsf{http://www.jalview.org/examples/uniref50.fa}).
+%
+% Are there differences ? If not, why not ?
+% }
+% }
+
+\section{Protein Alignment Conservation Analysis}
+\label{aacons}
+The {\sl Web Service $\Rightarrow$ Conservation} menu controls the computation
+of up to 17 different amino acid conservation measures for the current alignment
+view. The JABAWS AACon Alignment Conservation Calculation Service, which is used
+to calculate these scores, provides a variety of standard measures described by
+Valdar in 2002\footnote{Scoring residue conservation. Valdar (2002) {\sl
+Proteins: Structure, Function, and Genetics} {\bf 43} 227-241.} as well as an efficient implementation of the SMERFs
+score developed by Manning et al. in 2008.\footnote{SMERFS Score Manning et al. {\sl BMC
+Bioinformatics} 2008, {\bf 9} 51 \href{http://dx.doi.org/10.1186/1471-2105-9-51}{doi:10.1186/1471-2105-9-51}}
+
+\subsubsection{Enabling and Disabling AACon Calculations}
+When the AACon Calculation entry in the {\sl Web Services $\Rightarrow$
+Conservation} menu is ticked, AACon calculations will be performed every time
+the alignment is modified. Selecting the menu item will enable or disable
+automatic recalculation.
+
+\subsubsection{Configuring which AACon Calculations are Performed}
+The {\sl Web Services $\Rightarrow$ Conservation $\Rightarrow$ Change AACon
+Settings ...} menu entry will open a web services parameter dialog for the
+currently configured AACon server. Standard presets are provided for quick and
+more expensive conservation calculations, and parameters are also provided to
+change the way that SMERFS calculations are performed.
+AACon settings for an alignment are saved in Jalview projects along with the
+latest calculation results.
+
+\subsubsection{Changing the Server used for AACon Calculations}
+If you are working with alignments too large to analyse with the public JABAWS
+server, then you will most likely have already configured additional JABAWS
+servers. By default, Jalview will chose the first AACon service available from
+the list of JABAWS servers available. If available, you can switch to use
+another AACon service by selecting it from the {\sl Web Services $\Rightarrow$
+Conservation $\Rightarrow$ Switch Server} submenu.
+
+\chapter{Analysis of Alignments}
+\label{alignanalysis}
+Jalview provides support for sequence analysis in two ways. A number of
+analytical methods are `built-in', these are accessed from the {\sl Calculate}
+alignment window menu. Computationally intensive analyses are run outside
+Jalview {\sl via} web services - these are typically accessed {\sl via} the {\sl
+Web Service} menu, and described in chapter \ref{jvwebservices}.
+In this section, we describe the built-in analysis capabilities common to both
+the Jalview Desktop and the JalviewLite applet.
+
+\section{PCA}
+This calculation creates a spatial representation of the similarities within the
+current selection or the whole alignment if no selection has been made. After
+the calculation finishes, a 3D viewer displays the each sequence as a point in
+3D `similarity space'. Sets of similar sequences tend to lie near each other in
+this space.
Note: The calculation is computationally expensive, and may fail for very large
sets of sequences - because the JVM has run out of memory. Memory issues, and
how to overcome them, were discussed in Section \ref{memorylimits}.
\subsubsection{What is PCA?}
Principal components analysis is a technique for examining the structure of
complex data sets. The components are a set of dimensions formed from the
-measured values in the data set, and the principle component is the one with the
+measured values in the data set, and the principal component is the one with the
greatest magnitude, or length. The sets of measurements that differ the most
-should lie at either end of this principle axis, and the other axes correspond
+should lie at either end of this principal axis, and the other axes correspond
to less extreme patterns of variation in the data set.
In this case, the components are generated by an eigenvector decomposition of
the matrix formed from the sum of pairwise substitution scores at each aligned
in Jalview's built in documentation.\footnote{See
\url{http://www.jalview.org/help/html/calculations/pca.html}.}
+
+\exercise{Principal Component Analysis}{ \exstep{Load the alignment at
+\textsf{http://www.jalview.org/tutorial/alignment.fa} }
+\exstep{Select the menu option {\sl Calculate $\Rightarrow$ Principle Component Analysis}.
+A new window will open. Move this window so that the tree, alignment and PCA viewer window are all visible.
+Try rotating the plot by clicking and dragging the mouse on the plot in the PCA window.
+Note that clicking on points in the plot will highlight them on the alignment. }
+\exstep{ Select {\sl Calculate $\Rightarrow$ Calculate Tree $\Rightarrow$
+Neighbour Joining Using BLOSUM62}. A new tree window will appear.
+Click on the tree window. Careful selection of the tree partition location will divide the alignment into a number of groups,
+each of a different colour.
+Note how the colour of the sequence ID label matches both the colour of
+the partitioned tree and the points in the PCA plot.} }
+
\subsubsection{The PCA Viewer}
-PCA analysis can be launched from the {\sl Calculate $\Rightarrow$ Principle
+PCA analysis can be launched from the {\sl Calculate $\Rightarrow$ Principlal
Component Analysis} menu option. {\bf PCA requires a selection containing at
least 4 sequences}. A window opens containing the PCA tool (Figure \ref{PCA}).
Each sequence is represented by a small square, coloured by the background
keys or the scroll wheel of the mouse (if available). A tool tip appears if the
cursor is placed over a sequence. Sequences can be selected by clicking on them.
[CTRL]-Click can be used to select multiple sequences.
+
+Labels will be shown for each sequence by toggling the {\sl View $\Rightarrow$
+Show Labels} menu option, and the plot background colour changed {\sl via} the
+{\sl View $\Rightarrow$ Background Colour..} dialog box. A graphical
+representation of the PCA plot can be exported as an EPS or PNG image {\sl via}
+the {\sl File $\Rightarrow$ Save As $\Rightarrow$ \ldots } submenu.
+
\begin{figure}[hbtp]
\begin{center}
\includegraphics[width=2in]{images/PCA1.pdf}
\includegraphics[width=3in]{images/PCA3.pdf}
-\caption{{\bf PCA Analysis} }
+\caption{{\bf PCA Analysis.} }
\label{PCA}
\end{center}
\end{figure}
-Labels will be shown for each sequence by toggling the {\sl View $\Rightarrow$
-Show Labels} menu option, and the plot background colour changed {\sl via} the
-{\sl View $\Rightarrow$ Background Colour..} dialog box. A graphical
-representation of the PCA plot can be exported as an EPS or PNG image {\sl via}
-the {\sl File $\Rightarrow$ Save As $\Rightarrow$ \ldots } submenu.
-\exercise{Principle Component Analysis}{ \exstep{Load the alignment at
-\textsf{http://www.jalview.org/examples/exampleFile.jar} and press [ESC] to clear any selections. Alternatively, select {\sl Select $\Rightarrow$ Undefine Groups} to remove all groups and colourschemes. } \exstep{Select the menu option {\sl Calculate $\Rightarrow$ Principle Component Analysis}. A new window will open. Move this window so that the tree, alignment and PCA viewer window are all visible. Try rotating the plot by clicking and dragging the mouse on the plot in the PCA window. Note that clicking on points in the plot will highlight them on the alignment and tree. }
-\exstep{ Click on the tree window. Careful selection of the tree partition
-location will divide the alignment into a number of groups, each of a different
-colour. Note how the colour of the sequence ID label matches both the colour of
-the partitioned tree and the points in the PCA plot.
-} }
-\subsubsection{PCA data export}
+\subsubsection{PCA Data Export}
Although the PCA viewer supports export of the current view, the plots produced
are rarely suitable for direct publication. The PCA viewer's {\sl File} menu
includes a number of options for exporting the PCA matrix and transformed points
as comma separated value (CSV) files. These files can be imported by tools such
as {\bf R} or {\bf gnuplot} in order to graph the data.
-\subsection{Trees}
+\section{Trees}
\label{trees}
Jalview can calculate and display trees, providing interactive tree-based
grouping of sequences though a tree viewer. All trees are calculated {\sl via}
\includegraphics[width=2.5in]{images/trees1.pdf}
\includegraphics[width=2.5in]{images/trees2.pdf}
\includegraphics[width=1.25in]{images/trees4.pdf}
-\caption{{\bf Calculating Trees} Jalview provides four built in models for calculating trees. Jalview can also load precalculated trees in Newick format (right).}
+\caption{{\bf Calculating Trees} Jalview provides four built in models for calculating trees.
+Jalview can also load precalculated trees in Newick format (right).}
\label{trees1}
\end{center}
\end{figure}
\begin{figure}
\begin{center}
\includegraphics[width=5in]{images/trees3.pdf}
-\caption{{\bf Interactive Trees} The tree level cutoff can be used to designate groups in Jalview}
+\caption{{\bf Interactive Trees} The tree level cutoff can be used to designate
+groups in Jalview.}
\label{trees2}
\end{center}
\end{figure}
% move to ch. 3 ?
%Both PCA and Tree viewers are linked analysis windows. This means that their selection and display are linked to a particular alignment, and control and reflect the selection state for a particular view.
-\subsubsection{Recovering input data for a tree or PCA plot calculation}
+\subsubsection{Recovering input Data for a Tree or PCA Plot Calculation}
\parbox[c]{5in}{
The {\sl File $\Rightarrow$ Input Data } option will open a new alignment window containing the original data used to calculate the tree or PCA plot (if available). This function is useful when a tree has been created and then the alignment subsequently changed.
}
\parbox[c]{1.25in}{\centerline{\includegraphics[width=1.25in]{images/pca_fmenu.pdf}
}}
-\subsubsection{Changing the associated view for a tree or PCA viewer}
+\subsubsection{Changing the associated View for a Tree or PCA Viewer}
\parbox[c]{4in}{
The {\sl View $\Rightarrow$ Associated Nodes With $\Rightarrow$ .. } submenu is shown when the viewer is associated with an alignment that is involved in multiple views. Selecting a different view does not affect the tree or PCA data, but will change the colouring and display of selected sequences in the display according to the colouring and selection state of the newly associated view.
} \parbox[c]{3in}{\centerline{
\includegraphics[width=2.5in]{images/pca_vmenu.pdf} }}
-
-\exercise{Trees}{
-\exstep{Ensure that you have at least 1G memory available in Jalview (start with this link: \href{http://www.jalview.org/webstart/jalview_1G.jnlp}{http://www.jalview.org/webstart/jalview\_1G.jnlp}).}
-\exstep{Open the alignment at \textsf{http://www.jalview.org/tutorial/alignment.fa}. Select {\sl Calculate $\Rightarrow$ Calculate Tree $\Rightarrow$ Neighbour Joining Using BLOSUM62}. A new tree window will appear.}
-\exstep{Click on the tree window. A cursor will appear. Note that placing this cursor divides the tree into a number of groups by colour. Place the cursor to give about 4 groups, then select {\sl Calculate $\Rightarrow$ Sort $\Rightarrow$ By Tree Order $\Rightarrow$ Neighbour Joining Tree using BLOSUM62 from ... }. The sequences are reordered to match the order in the tree and groups are formed implicitly.}
-\exstep{Select {\sl Calculate $\Rightarrow$ Calculate Tree $\Rightarrow$ Neighbour Joining Using \% Identity}. A new tree window will appear. The group colouring makes it easy to see the diferences between the two trees, calculated using different methods.}
-\exstep{Select from sequence 2 column 60 to sequence 12 column 123. Select {\sl Calculate $\Rightarrow$ Calculate Tree $\Rightarrow$ Neighbour Joining Using BLOSUM62}. A new tree window will appear. It can be seen that the tree contains 11 sequences. It has been coloured according to the already selected groups from the first tree and is calculated purely from the residues in the selection.
-Comparing the location of individual sequences between the three trees illustrates the importance of selecting appropriate regions of the alignment for the calculation of trees.
-}
-\exstep{Recover the {\sl Input Data} for the tree you just calculated from the {\sl File} menu. Check the {\sl Edit $\Rightarrow$ Pad Gaps } option is {\sl not} ticked, and insert one gap anywhere in the alignment. Now select {\sl Calculate $\Rightarrow$ Calculate Tree $\Rightarrow$ Neighbour Joining Using BLOSUM62}.
-
-A warning dialog box {\bf ``Sequences not aligned'' } appears because the sequences input to the tree calculation are of different lengths. }
-
-\exstep{Now select {\sl Edit $\Rightarrow$ Pad Gaps } and try to perform the tree calculation again - this time a new tree should appear.
-
-This demonstrates the use of the {\sl Pad Gaps } editing preference, which ensures that all sequences are the same length after editing. }
-
-}
-
-\subsection{Tree Based Conservation Analysis}
-\label{treeconsanaly}
+\subsection{Tree Based Conservation Analysis}
+\label{treeconsanaly}
Trees reflect the pattern of global sequence similarity exhibited by the
alignment, or region within the alignment, that was used for their calculation.
Annotation $\Rightarrow$ Group Conservation} and {\sl Group Consensus} options)
can help when working with larger alignments.
+\exercise{Trees}{
+\exstep{Ensure that you have at least 1G memory available in Jalview
+(Either start with this link:
+\href{http://www.jalview.org/services/launchApp?jvm-max-heap=1G}{http://www.jalview.org/services/launchApp?jvm-max-heap=1G},
+or in the Development section of the Jalview web site
+(\href{http://www.jalview.org/development/development-builds}{http://www.jalview.org/development/development-builds})
+in the ``latest official build'' row in the table, go to the
+``Webstart'' column, click on ``G2''.)}
+\exstep{Open the alignment at \textsf{http://www.jalview.org/tutorial/alignment.fa}.
+Select {\sl Calculate $\Rightarrow$ Calculate Tree $\Rightarrow$ Neighbour Joining Using BLOSUM62}. A new tree window will appear.}
+\exstep{Click on the tree window. A cursor will appear. Note that placing this cursor divides the tree into a number of groups by colour. Place the cursor to give about 4 groups, then select {\sl Calculate $\Rightarrow$ Sort $\Rightarrow$ By Tree Order $\Rightarrow$ Neighbour Joining Tree using BLOSUM62 from ... }. The sequences are reordered to match the order in the tree and groups are formed implicitly.}
+\exstep{Select {\sl Calculate $\Rightarrow$ Calculate Tree $\Rightarrow$
+Neighbour Joining Using \% Identity}. A new tree window will appear. The group colouring
+makes it easy to see the differences between the two trees, calculated using
+ different methods.} \exstep{Select from sequence 2 column 60 to sequence 12 column 123. Select {\sl Calculate $\Rightarrow$ Calculate Tree $\Rightarrow$ Neighbour Joining Using BLOSUM62}. A new tree window will appear. It can be seen that the tree contains 11 sequences. It has been coloured according to the already selected groups from the first tree and is calculated purely from the residues in the selection.
+Comparing the location of individual sequences between the three trees illustrates the importance of selecting appropriate regions of the alignment for the calculation of trees.
+}
+\exstep{Recover the {\sl Input Data} for the tree you just calculated from the {\sl File} menu. Check the {\sl Edit $\Rightarrow$ Pad Gaps } option is {\sl not} ticked, and insert one gap anywhere in the alignment. Now select {\sl Calculate $\Rightarrow$ Calculate Tree $\Rightarrow$ Neighbour Joining Using BLOSUM62}.
+
+A warning dialog box {\bf ``Sequences not aligned'' } appears because the sequences input to the tree calculation are of different lengths. }
+
+\exstep{Now select {\sl Edit $\Rightarrow$ Pad Gaps } and try to perform the tree calculation again - this time a new tree should appear.
+
+This demonstrates the use of the {\sl Pad Gaps } editing preference, which ensures that all sequences are the same length after editing. }
+
+}
+
\exercise{Tree Based Conservation Analysis}{
\label{consanalyexerc}
\exstep{Load the PF03460 PFAM seed alignment using the sequence fetcher. Colour it with the {\sl Taylor colourscheme}, and apply {\sl Conservation } shading. }
-\exstep{Build a Neighbourjoining tree using BLOSUM62 and use the {\sl Sort Alignment By Tree} option in the tree viewer submenu to order alignment using the calculated tree.}
-\exstep{Select a point on the tree to partition the alignment, and examine the variation in colouring between different groups.
+\exstep{Build a Neighbour joining tree using BLOSUM62 and use the {\sl Sort
+Alignment By Tree} option in the tree viewer submenu to order alignment using the calculated tree.} \exstep{Select a point on the tree to partition the alignment, and examine the variation in colouring between different groups.
-You may find it easier to browse the alignment if you first uncheck the {\sl View $\Rightarrow$ Show Annotations} option, and open the Overview Window to aid navigation.}
+You may find it easier to browse the alignment if you first uncheck the {\sl
+Annotations $\Rightarrow$ Show Annotations} option, and open the Overview Window
+within the View menu to aid navigation.}
\exstep{Try changing the colourscheme to BLOSUM62 (whilst ensuring that {\sl Apply Colour to All Groups} is selected)}
{\sl Note: You may want to save the alignment and tree as a project file, since
it is used in the next few exercises. } }
+
\subsection{Redundancy Removal}
The redundancy removal dialog box is opened using the {\sl Edit $\Rightarrow$ Remove Redundancy\ldots} option in the alignment menu. As its menu option placement suggests, this is actually an alignment editing function, but it is convenient to describe it here. The redundancy removal dialog box presents a percentage identity slider which sets the redundancy threshold. Aligned sequences which exhibit a percentage identity greater than the current threshold are highlighted in black. The [Remove] button can then be used to delete these sequences from the alignment as an edit operation\footnote{Which can usually be undone. A future version of Jalview may allow redundant sequences to be hidden, or represented by a chosen sequence, rather than deleted.}.
\caption{The Redundancy Removal dialog box opened from the edit menu. Sequences that exceed the current percentage identity threshold and are to be removed are highlighted in black.}
\end{figure}
-\exercise{Remove redundant sequences}{
-
-{\sl Note: Jalview 2.8 users - bugs in this version mean that the 'Unlinked leaves' markings will not be shown when sequences are removed during this exercise.}
-
-\exstep{Re-use or recreate the alignment and tree which you worked with in the
-tree based conservation analysis exercise (exercise \ref{consanalyexerc})}
-\exstep{Open the Remove Redundancy dialog and adjust the threshold to 90\%. Remove the sequences that are more than 90\% similar under this alignment.}
-\exstep{Select the Tree viewer's {\sl View $\Rightarrow$ Mark Unlinked Leaves} option, and note that the removed sequences are now prefixed with a * in the tree view.}
-\exstep{Use the [Undo] button on the dialog to recover the sequences. Note that the * symbols disappear from the tree display.}
-\exstep{Experiment with the redundancy removal and observe the relationship between the percentage identity threshold and the pattern of unlinked nodes in the tree display.}
-}
-\subsection{Subdividing the alignment according to specific mutations}
+\subsection{Subdividing the Alignment According to Specific Mutations}
It is often necessary to explore variations in an alignment that may correlate
with mutations observed in a particular region; for example, sites exhibiting
colourschemes can then be used to reveal any associated pattern of sequence
variation across the whole alignment.
-\subsection{Automated annotation of Alignments and Groups}
-On loading a sequence alignment, Jalview will normally\footnote{Automatic
-annotation can be turned off in the {\sl Visual } tab in the {\sl Tools
-$\Rightarrow$ Preferences } dialog box.} calculate a set of automatic annotation
-rows which are shown below the alignment. For nucleotide sequence alignments,
-only an alignment consensus row will be shown, but for amino acid sequences,
-alignment quality (based on BLOSUM 62) and physicochemical conservation will
-also be shown. Conservation is calculated according to Livingstone and
-Barton\footnote{{\sl ``Protein Sequence Alignments: A Strategy for the
-Hierarchical Analysis of Residue Conservation." } Livingstone C.D. and Barton
-G.J. (1993) {\sl CABIOS } {\bf 9}, 745-756}.
-Consensus is the modal residue (or {\tt +} where there is an equal top residue).
-The inclusion of gaps in the consensus calculation can be toggled by
-right-clicking on the the Consensus label and selecting {\sl Ignore Gaps in
-Consensus} from the context menu. Quality is a measure of the inverse likelihood
-of unfavourable mutations in the alignment. Further details on these
-calculations can be found in the on-line documentation.
+% These annotations can be hidden and deleted via the context menu linked to the
+% annotation row; but they are only created on loading an alignment. If they are
+% deleted then the alignment should be saved and then reloaded to restore them.
+% Jalview provides a toggle to autocalculate a consensus sequence upon editing.
+% This is normally selected by default, but can be turned off for large alignments {\sl via} the {\sl Calculate $\Rightarrow$ Autocalculate
+% Consensus} menu option if the interface is too slow.
-These annotations can be hidden and deleted but are only created on loading an
-alignment. If they are deleted then the alignment should be saved and reloaded
-to restore them. Jalview provides a toggle to autocalculate a consensus sequence
-upon editing. This is normally selected by default, but can be turned off for
-large alignments {\sl via} the {\sl Calculate $\Rightarrow$ Autocalculate
-Consensus} menu option if the interface is too slow.
+% \subsubsection{Group Associated Annotation}
+% \label{groupassocannotation}
+% Group associated consensus and conservation annotation rows reflect the
+% sequence variation within a particular group. Their calculation is enabled
+% by selecting the {\sl Group Conservation} or {\sl Group Consensus} options in
+% the {\sl Annotation $\Rightarrow$ Autocalculated Annotation } submenu of the
+% alignment window.
-\subsubsection{Group Associated Annotation}
-\label{groupassocannotation}
-Group associated consensus and conservation annotation rows reflect the
-sequence variation within a particular group. Their calculation is enabled
-by selecting the {\sl Group Conservation} or {\sl Group Consensus} options in
-the {\sl View $\Rightarrow$ Autocalculated Annotation } submenu of the alignment
-window.
+% \subsubsection{Alignment and Group Sequence Logos}
+% \label{seqlogos}
-\subsubsection{Alignment and Group Sequence Logos}
-\label{seqlogos}
+% The consensus annotation row that is shown below the alignment can be overlaid
+% with a sequence logo that reflects the symbol distribution at each column of
+% the alignment. Right click on the Consensus annotation row and select the {\sl
+% Show Logo} option to display the Consensus profile for the group or alignment.
+% Sequence logos can be enabled by default for all new alignments {\sl via} the
+% Visual tab in the Jalview desktop's preferences dialog box.
-The consensus annotation row that is shown below the alignment can be overlaid
-with a sequence logo that reflects the symbol distribution at each column of
-the alignment. Right click on the Consensus annotation row and select the {\sl Show
-Logo} option to display the Consensus profile for the group or alignment.
-Sequence logos can be enabled by default for all new alignments {\sl via} the
-Visual tab in the Jalview desktop's preferences dialog box.
+\section{Pairwise Alignments}
+Jalview can calculate optimal pairwise alignments between arbitrary
+sequences {\sl via} the {\sl Calculate $\Rightarrow$ Pairwise Alignments\ldots} menu option.
+Global alignments of all pairwise combinations of the selected sequences are performed and the results returned in a text box.
+
+
+
+\exercise{Remove Redundant Sequences}{
+
+\exstep{Re-use or recreate the alignment and tree which you worked with in the
+tree based conservation analysis exercise (exercise \ref{consanalyexerc}). In
+the alignment window, you may need to deselect groups using Esc key.}
+\exstep{In the Edit menu select Remove Redundancy to open the Redundancy
+threshold selection dialog. Adjust the redundancy threshold value, start
+at 50 and increase the value to 65. Sequences selected will change colour in the Sequence ID panel. Select ``Remove'' to
+remove the sequences that are more than 65\% similar under this alignment.}
+\exstep{Select the Tree viewer's {\sl View $\Rightarrow$ Mark Unlinked Leaves} option, and note that the removed sequences are now prefixed with a * in the tree view.}
+\exstep{Use the [Undo] button in the Redundancy threshold selection dialog box
+to recover the sequences. Note that the * symbols disappear from the tree display.}
+\exstep{Experiment with the redundancy removal and observe the relationship between the percentage identity threshold and the pattern of unlinked nodes in the tree display.}
+}
-\exercise{Group conservation analysis}{
+\exercise{Group Conservation Analysis}{
\exstep{Re-use or recreate the alignment and tree which you worked with in the
tree based conservation analysis exercise (exercise \ref{consanalyexerc})}
-\exstep{Create a new view, and ensure the annotation panel is displayed, and
-enable the display of {\sl Group Consensus} and the display of sequence
-logos to make it easier to see the different residue populations within each group.}
-\exstep{Select a column exhibiting about 50\% conservation that lies within the
-central conserved region of the alignment. Subdivide the alignment according to
+\exstep{In the View menu, create a new view. Ensure the annotation panel
+is displayed (Show annotation in Annotations menu). Enable the display
+of {\sl Group Consensus} option by checking {\sl Group Consensus} in the {\sl Annotation $\Rightarrow$
+Autocalculated Annotation } submenu in the sequence alignment window. Then display of sequence
+logos to make it easier to see the different residue populations within each
+group. Activate logo by right clicking on the Consensus annotation row to open
+the pop-up menu and select the {\sl Show Logo} option.}
+\exstep{In the column alignment ruler, select a column exhibiting about 50\%
+conservation that lies within the central conserved region of the alignment. Subdivide the alignment according to
this selection using {\sl Select $\Rightarrow$ Make groups for selection}.}
\exstep{Re-order the alignment according to the new groups that have been
defined. Click on the group annotation row IDs to select groups exhibiting a
the tree groups made in the previous exercise.}
}
-\subsection{Other Calculations}
-
-
-\subsubsection{Pairwise Alignments}
-
-Jalview can calculate optimal pairwise alignments between arbitrary sequences {\sl via} the {\sl Calculate $\Rightarrow$ Pairwise Alignments\ldots} menu option. Global alignments of all pairwise combinations of the selected sequences are performed and the results returned in a text box.
-
\begin{figure}[]
\begin{center}
\includegraphics[width=4in]{images/pairwise.pdf}
\end{center}
\end{figure}
-\pagebreak[2]
-
-\section{Webservices}
-\label{jvwebservices}
-The term ``Webservices'' refers to a variety of data exchange
-mechanisms based on HTTP.\footnote{HTTP: Hyper-Text Transfer Protocol.}
-
-\parbox[c]{4.5in}{Jalview can exploit public webservices to access databases
-remotely, and also submit data to public services by opening pages with your web browser. These types of
-services are `one-way', {\sl i.e.} data is either sent to the webservice or
-retrieved from it by Jalview. The desktop application can also interact
-with `two-way' remote analysis services in order to offload computationally
-intensive tasks to High Performance Computing facilities. Most of these two-way
-services are provided by {\bf Ja}va {\bf B}ioinformatics {\bf A}nalysis {\bf
-W}eb {\bf S}ervice (JABAWS) servers\footnote{See
-http://www.compbio.dundee.ac.uk/jabaws for more information and to download
-your own server.}, which provides an easily installable system for performing
-a range of bioinformatics analysis tasks. }
-\parbox[c]{1.75in}{\includegraphics[width=1.65in]{images/wsmenu.pdf}}
-
-\subsection{One-way web services}
-
-There are three types of one way service in jalview. Database services,
-which were introduced in in Section \ref{fetchseq}, provide sequence and
-alignment data. They can also be used to add sequence IDs to an alignment
-imported from a local file, prior to further annotation retrieval, as described
-in Section \ref{featuresfromdb}. A second type of one way service is provided
-by Jalview's DAS sequence feature retrieval system, which is described
-in Section \ref{dasfretrieval}. The final type of one way service are sequence
-and ID submission services, exemplified by the `Envision2 Services' provided
-by the ENFIN Consortium\footnote{ENFIN is the European Network for Functional
-INtegration. Please see http://www.enfin.org for more information. }.
-
-\subsubsection{One-way submission services}
-Jalview can use the system's web browser to submit sets of sequences and
-sequence IDs to web based applications. Single sequence IDs can be passed to
-a web site using the user definable URL links listed under the {\sl
-Links} submenu of the sequence ID popup menu. These are configured
-in the {\sl Connections} tab of the {\sl Preferences} dialog box.
-
-The Envision 2 services presented in the webservice menu provides are the first
-example of one-way services where multiple sequences or sequence IDs can be
-sent. The {\sl Web service $\Rightarrow$ Envision 2 Services} menu entry
-provides two sub-menus that enable you to submit the sequences or IDs
-associated with the alignment or just the currently selected sequences to one
-of the Envision2 workflows. Selecting any one will open a new browser window on
-the Envision2 web application. The menu entries and their tooltips provide
-details of the Envision2 workflow and the dataset set that will be submitted
-({\sl i.e.} the database reference type, or associated sequence subset). Please
-note, due to technical limitations, Jalview can currently only submit small
-numbers of sequences to the workflows - if no sequence or ID submissions are
-presented in the submenus, then try to select a smaller number of sequences to
-submit.
-
-\subsection{Remote Analysis Web Services}
-Remote analysis services enable Jalview to use external computational
-facilities. There are curently three types of service - multiple sequence
-alignment, protein secondary structure prediction, and alignment analysis.
-Many of these are provided by JABA servers, which are described at the end of
-this section. In all cases, Jalview will construct a job based on the alignment
-or currently selected sequences, ask the remote server to run the job, monitor
-status of the job and, finally, retrieve the results of the job and display
-them. The Jalview user is kept informed of the progress of the job through a
-status window.
-Currently, web service jobs and their status windows are not stored in Jalview
-Project Files\footnote{This may be rectified in future versions.}, so it is
-important that you do not close Jalview whilst a job is running. It is also
-essential that you have a continuous network connection in order to
-successfully use web services from Jalview, since it periodically checks the
-progress of running jobs.
+\chapter{Working with 3D structures}
+\label{3Dstructure}
-\subsection{JABA Web Services for sequence alignment and analysis}
-\label{jabaservices}
-JABA stands for ``JAva Bioinformatics Analysis'', which is a system developed
-by Peter Troshin and Geoff Barton at the University of Dundee for running
-computationally intensive bioinformatics analysis programs. A JABA installation
-typically provides a range of JABA web services (JABAWS) for use by other
-programs, such as Jalview.
+To summarise, section \ref{featannot} describes the mechanisms provided by
+Jalview for interactive creation of sequence and alignment annotation, and how they can be
+displayed, imported and exported and used to reorder the alignment. Section
+\ref{featuresfromdb} discusses the retrieval of database references and
+establishment of sequence coordinate systems for the retrieval and display of
+features from databases and DAS annotation services.
+Section \ref{msaservices} describes how to use the range of multiple alignment
+programs provided by JABAWS, and Section \ref{aacons} introduces JABAWS AACon
+service for protein multiple alignment conservation analysis.
+ In Section \ref{alignanalysis}, you will find
+descriptions and exercises on building and displaying trees, PCA analysis,
+alignment redundancy removal, pairwise alignments and alignment conservation
+analysis.
+Section \ref{wkwithstructure} introduces the structure visualization
+capabilities of Jalview.
+Section \ref{protsspredservices} explains how to perform protein secondary
+structure predictions with JPred, and JABAWS protein disorder prediction
+services are introduced in Section \ref{protdisorderpred}.
+Section \ref{workingwithnuc} describes functions and visualization techniques relevant
+to working with nucleotide sequences, coding region annotation and nucleotide
+sequence alignments.
+Section \ref{jvwebservices} introduces the various web based services
+available to Jalview users, and Section \ref{jabaservices} explains how to
+configure the Jalview Desktop for access to new JABAWS servers.
-Exercises in the remainder of this section will demonstrate the simplest way of
-installing JABA on your computer, and configuring Jalview so it can access the JABA services. If you
-need any further help or more information about the services, please go to the
-\href{http://www.compbio.dundee.ac.uk/jabaws}{JABAWS home page}.
-%% \subsubsection{Aims}
-%% \begin{list}{$\bullet$}{}
-%% \item Gain experience using the different alignment services provided by
-% JABA
-%%\item Learn about the way that Jalview stores user presets for JABA services
-%%\item Learn how to install JABA services and configure Jalview to access them
-%%\end{list}
+
+% and Section \ref{workingwithrna} covers the visualization,
+% editing and analysis of RNA secondary structure.
-\subsection{Changing the Web Services menu layout}
-\label{changewsmenulayout}
-If you are working with a lot of different JABA services, you may wish to change
-the way Jalview lays out the web services menu. You can do this from the Web
-Services tab of the {\sl Preferences} dialog box.
+\section{Working with Structures}
+\label{wkwithstructure}
+Jalview facilitates the use of protein structures for the analysis of alignments
+by providing a linked view of structures associated with sequences in
+the alignment. The Java based molecular viewing program Jmol\footnote{See the
+Jmol homepage \url{http://www.jmol.org} for more information.} has been
+incorporated\footnote{Earlier versions of Jalview included MCView - a simple
+main chain structure viewer. Structures are visualized as an alpha carbon trace
+and can be viewed, rotated and coloured in a structure viewer and the results
+interpreted on a sequence alignment.} which enables sophisticated molecular
+visualizations to be prepared and investigated alongside an analysis of
+associated sequences.
+PDB format files can be imported directly or structures can be retrieved from
+the European Protein Databank (PDBe) using the Sequence Fetcher (see
+\ref{fetchseq}).
-\exercise{Changing the Layout of the Web Services Menu}{
-\label{changewsmenulayoutex}
-\exstep{Make sure you have loaded an alignment into Jalview, and examine the
-current layout of the alignment windowÔøΩs {\sl Web Service} menu.}
-\exstep{Open the preferences dialog box and select the web services tab.}
-\exstep{Ensure the {\sl Enable JABAWS services} checkbox is selected, and unselect
-the {\sl Enable Enfin Services} checkboxes.}
-\exstep{Hit {\sl Refresh Services} to update the web services menu -- once the
-progress bar has completed, open the {\sl Web Service} menu to view the changes.}
-\exstep{Select the {\sl Index by host} checkbox and refresh the services once again.
+\subsection{Automatic Association of PDB Structures with Sequences}
+Jalview can automatically determine which structures are associated with a
+sequence in a number of ways.
+\subsubsection{Discovery of PDB IDs from Sequence Database Cross-references}
+If a sequence has an ID from a public database that contains cross-references to
+the PDB, such as Uniprot. Right-click on any sequence ID and select {\sl Structure $\Rightarrow$
+Associate Structure with Sequence $\Rightarrow$ Discover PDB IDs } from the context menu (Figure \ref{auto}). Jalview will attempt to associate the
+sequence with a Uniprot sequence and from there discover any associated PDB
+structures. This takes a few seconds and applies to all sequences in the
+alignment which have valid Uniprot IDs. On moving the cursor over the sequence
+ID the tool tip\footnote{Tip: The sequence ID tooltip can often become large for
+heavily cross referenced sequence IDs. Use the {\sl View $\Rightarrow$ Sequence
+ID Tooltip $\Rightarrow$ } submenu to disable the display of database cross
+references or non-positional features. } now shows the Uniprot ID and any
+associated PDB structures.
-{\sl Observe the way the layout of the JABAWS Alignment submenu changes.}
+\begin{figure}[htbp]
+\begin{center}
+%TODO fix formatting
+\parbox{1.5in}{
+{\centering
+\begin{center}
+\includegraphics[width=1.5in]{images/auto1.pdf}
+\end{center}}
+} \parbox{3.25in}{
+{\centering
+\begin{center}
+\includegraphics[scale=0.5]{images/auto2.pdf}
+\end{center}
+}
+} \parbox{1.5in}{
+{\centering
+\begin{center}
+\includegraphics[width=1.5in]{images/auto3.pdf}
+\end{center}
}
-\exstep{Do the same with the {\sl Index by type} checkbox.}
}
-Jalview provides these options for configuring the layout of the {\sl Web Service}
-menu because different Jalview users may have access to a different number of
-JABA services, and each will have their own preference regarding the layout of
-the menu.
-
-\begin{figure}[htbc]
-\begin{center}
-\includegraphics[width=3in]{images/jvjabawsconfig.pdf}
-\caption{{\bf The Jalview Web Services preferences panel.} Options are provided
-for configuring the list of JABA servers that Jalview will use, enabling and
-disabling Enfin services, and configuring the layout of the web services
-menu.}
-\label{jvjabawsconfig}
+\caption{{\bf Automatic PDB ID discovery.} The tooltip (left) indicates that no PDB structure has been associated with the sequence.
+After PDB ID discovery (center) the tool tip now indicates the Uniprot ID and
+any associated PDB structures (right).}
+\label{auto}
\end{center}
\end{figure}
+\subsubsection{Drag-and-Drop Association of PDB Files with Sequences by Filename
+Match}
+\label{multipdbfileassoc}
+If one or more PDB files stored on your computer are dragged from their location
+on the file browser onto an alignment window, Jalview will search the alignment
+for sequences with IDs that match any of the files dropped onto the alignment.
+If it discovers matches, a dialog like the one in Figure
+\ref{multipdbfileassocfig} is shown, giving the option of creating associations
+for the matches.
-\subsubsection{Testing JABA services}
-The JABAWS configuration dialog shown in Figure \ref{jvjabawsconfig} has colour
-codes to indicate whether the Desktop can access the server, and whether all
-services advertised by the server are functional. The colour codes are:
-
-\begin{list}{$\bullet$}{}
- \item Red - Server cannot be contacted or reports a connection error.
- \item Amber - Jalview can connect, but one or more services are non-functional.
- \item Green - Server is functioning normally.
-\end{list}
- %TODO green and a tick, red and a cros, amber and a value indicating whether its all going
+If no associations are made, then sequences extracted
+from the structure will be simply added to the alignment. However, if only
+some of the PDB files are associated, Jalview will raise another dialog box
+giving you the option to add any remaining sequences from the PDB structure files not present in
+the alignment. This allows you to easily decorate sequences in a newly imported
+alignment with any corresponding structures you've already collected in a directory
+accessible from your computer.\footnote{We plan to extend this facility in
+future so Jalview will automatically search for PDB files matching your
+sequence within a local directory. Check out
+\href{http://issues.jalview.org/browse/JAL-801}{Jalview issue 801}}
-Test results from JABAWS are reported on Jalview's console output (opened from
-the Tools menu). Tests are re-run every time Jalview starts, and when the
-[Refresh] button is pressed on the Jalview JABAWS configuration panel.
+% there is no mention of the other footnote (#3) that appears saying: Tip: The sequence ID tooltip can often become large for heavily cross-referenced sequence IDs. Use the ...
+% JBP: yes there is - under 'Discovery of ' subsection.
+\begin{figure}[htbp]
+\begin{center}
+\includegraphics[]{images/pdbdragdropassoc.pdf}
-\subsubsection{Resetting the JABA services setting to their defaults}
-Once you have configured a JABAWS server and selected the OK button of the
-preferences menu, the settings will be stored in your Jalview preferences file,
-along with any preferences regarding the layout of the web services menu. If
-you should ever need to reset the JABAWS server list to its defaults, use the
-`Reset Services' button on the Web Services preferences panel.
-
-\subsection{Running your own JABA server}
-You can download and run JABA on your own machine using the `VMWare' or
-VirtualBox virtual machine environments. If you would like to learn how to do
-this, there are full instructions at the
-\href{http://www.compbio.dundee.ac.uk/jabaws/}{JABA web site}.
-
-\exercise{Installing a JABA Virtual Machine on your computer}{
-\label{jabawsvmex}{\sl This tutorial will demonstrate the simplest way of
-installing JABA on your computer, and configuring Jalview so it can access the JABA services.
+\caption{{\bf Associating PDB files with sequences by drag-and-drop.} Dragging
+PDB files onto an alignment of sequences with names matching the dragged files
+names (A), results in a dialog box (B) that gives the option to associate each
+file with any sequences with matching IDs. }
+\label{multipdbfileassocfig}
+\end{center}
+\end{figure}
-{\bf Prerequisites}
-You will need a copy of VMWare Player/Workstation/Fusion on your machine.
-}
+\subsection{Viewing Structures}
+\label{viewAllStructures}
+The structure viewer can be launched in two ways from the sequence ID context
+menu. To view a particular structure associated with a sequence in the
+alignment, simply select it from popup menu's associated structures submenu in
+{\sl Structure $\Rightarrow$ View Structure $\Rightarrow$ $<$PDB ID$>$}. The
+second way is most useful if you want to view all structural data available for
+a set of sequences in an alignment. If any of the {\bf currently selected}
+sequences have structures associated, the {\sl Structure } submenu of the
+sequence ID popup menu will include an option to {\sl View {\bf N}
+structures}. Selecting this option will open a new structure view containing
+the associated structures superposed according to the alignment.
-\exstep{If you do not have VMWare player installed, download it from
-www.vmware.com (this takes a few minutes -- you will need to register and wait
-for an email with a download link).}
-\exstep{Download the JABA virtual appliance archive called `jaba-vm.zip' from
-\textsf{http://www.compbio.dundee.ac.uk/jabaws/archive/jabaws-vm.zip}
+In both cases, each structure to be displayed will be downloaded or loaded from
+the local file system, and shown as a ribbon diagram coloured according to the
+associated sequence in the current alignment view (Figure \ref{structure}
+(right)). The structure can be rotated by clicking and dragging in the structure
+window. The structure can be zoomed using the mouse scroll wheel or by
+[SHIFT]-dragging the structure.
+Moving the mouse cursor over a sequence to which the structure is linked in the
+alignment view highlights the respective residue's sidechain atoms. The
+sidechain highlight may be obscured by other parts of the molecule. Similarly,
+moving the cursor over the structure shows a tooltip and highlights the
+corresponding residue in the alignment. Clicking the alpha carbon or phosphorous
+backbone atom will toggle the highlight and residue label on and off. Often, the
+position highlighted in the sequence may not be in the visible portion of the
+current alignment view and the sliders will scroll automatically to show the
+position. If the alignment window's {\sl View $\Rightarrow$ Automatic Scrolling
+} option is not selected, however, then the automatic adjustment will be
+disabled for the current view.
-WARNING: This is large (about 300MB) and will take some time to download.
+\begin{figure}[htbp]
+\begin{center}
+\parbox{3in}{
+{\centering
+\begin{center}
+\includegraphics[scale=0.5]{images/structure1.pdf}
+\end{center}
}
-\exstep{Unpack the archive's contents to a place on your machine with at least
-2GB of free space.
-
-(On Windows, right click on the archive, and use the 'Extract archive..' option).
}
-\exstep{Open the newly extracted directory and double click the VMWare virtual
-machine configuration file (jabaws.vcf). This will launch the VMWare player.
+\parbox{3.2in}{
+{\centering
+\begin{center}
+\includegraphics[width=3in]{images/structure2.pdf}
+\end{center}
}
-\exstep{Once VMWare player has started up, it may ask the question ``Did you move or copy
-this virtual appliance?'' -- select `Copy'.}
-\exstep{You may be prompted to download the VMWare linux tools. These are not
-necessary, so close the window or click on `Later'.}
-\exstep{You may also be prompted to install support for one or more devices (USB
-or otherwise). Say `No' to these options.}
-\exstep{Once the machine has loaded, it will display a series of IP addresses
-for the different services provided by the VM. Make a note of the JABAWS URL --
-this will begin with `http:' and end with `/jabaws''.}
}
+\caption{{\bf Structure visualization} The structure viewer is launched from the sequence ID context menu (left) and allows the structure to be visualized using the embedded Jmol molecular viewer (right). }
+\label{structure}
+\end{center}
+\end{figure}
-\exercise{Configuring Jalview to access your new JABAWS virtual appliance}{
-\label{confnewjabawsappl}
-\exstep{Start Jalview (If you have not done so already).}
-\exstep{Enable the Jalview Java Console by selecting its option from the Tools
-menu.
+\subsection{Customising Structure Display}
-{\sl Alternately, use the System Java console if you have
-configured it to open when Jalview is launched, {\sl via} your system's Java
-preferences (under the `Advanced' tab on Windows).}}
-\exstep{Open the {\sl Preferences} dialog and locate the Web Services tab.}
-\exstep{Add the URL for the new JABAWS server you started in Exercise
-\ref{jabawsvmex} to the list of JABAWS urls using the `New Service
-URL' button.}
-\exstep{You will be asked if you want to test the service. Hit `Yes' to do this
--- you should then see some output in the console window.
+Structure display can be modified using the {\sl Colour} and {\sl View} menus
+in the structure viewer. The background colour can be modified by selecting the
+{\sl Colours $\Rightarrow$ Background Colour\ldots} option.
-{\sl Take a close look at the output in the console. What do you think is
-happening?}
-}
-\exstep{Hit OK to save your preferences -- you have now added a new JABA
-service to Jalview!}
-\exstep{Try out your new JABA services by loading the ferredoxin sequences from
-http://www.jalview.org/tutorial/alignment.fa}
-\exstep{Launch an alignment using one
-of the JABA methods provided by your server. It will be listed under the JABAWS Alignment submenu of the {\sl Web Service} menu on the alignment window.
+By default, the structure will be coloured in the same way as the associated
+sequence(s) in the alignment view from which it was launched. The structure can
+be coloured independently of the sequence by selecting an appropriate colour
+scheme from the {\sl Colours} menu. It can be coloured according to the
+alignment using the {\sl Colours $\Rightarrow$ By Sequence } option. The image
+in the structure viewer can be saved as an EPS or PNG with the {\sl File
+$\Rightarrow$ Save As $\Rightarrow$ \ldots} submenu, which also allows the raw
+data to be saved as PDB format. The mapping between the structure and the
+sequence (How well and which parts of the structure relate to the sequence) can
+be viewed with the {\sl File $\Rightarrow$ View Mapping} menu option.
-{\sl Note: You can watch the JABA VM appliance's process working by opening the
-process monitor on your system. (On Windows XP, this involves right-clicking the
-system clock and opening the task manager -- then selecting the 'Processes' tab
-and sort by CPU).}
-}
+\subsubsection{Using the Jmol Visualization Interface }
+
+Jmol has a comprehensive set of selection and visualization functions that are
+accessed from the Jmol popup menu (by right-clicking in the Jmol window or by
+clicking the Jmol logo). Molecule colour and rendering style can be manipulated,
+and distance measurements and molecular surfaces can be added to the view. It
+also has its own ``Rasmol\footnote{See \url{http://www.rasmol.org}}-like'' scripting
+language, which is described elsewhere\footnote{Jmol Wiki:
+\url{http://wiki.jmol.org/index.php/Scripting}
+
+Jmol Scripting reference:
+\url{http://www.stolaf.edu/academics/chemapps/jmol/docs/}}. Jalview utilises the
+scripting language to interact with Jmol and to store the state of a Jmol
+visualization within Jalview archives, in addition to the PDB data file
+originally loaded or retrieved by Jalview. To access the Jmol scripting
+environment directly, use the {\sl Jmol $\Rightarrow$ Console} menu option.
+
+If you would prefer to use Jmol to manage structure colours, then select the
+{\sl Colours $\Rightarrow$ Colour with Jmol} option. This will disable any
+automatic application of colour schemes when new structure data is added, or
+when associated alignment views are modified.
+
+
+\exercise{Viewing Structures}{\label{viewingstructex}
+\exstep{Load the alignment at
+\textsf{http://www.jalview.org/examples/exampleFile.jar}. Right-click on the
+sequence ID label of {\sl FER1\_SPIOL}, this brings up
+the context menu. Select {\sl FER1\_SPIOL $\Rightarrow$ Structure $\Rightarrow$
+Associate Structure with Sequence $\Rightarrow$ Discover
+PDB IDs}. Jalview will now attempt to find PDB structures for the sequences in
+the alignment.
+% JBP Note: Bug JAL-1238 needs to be fixed ASAP
+
+{\bf Note:} If you are using Jalview v2.8 - use the {\sl Uniprot } source from the {\sl Web services $\Rightarrow$ Fetch DB References $\Rightarrow$ ..} submenu of the Alignment Window to retrieve the PDB IDs. }
+\exstep{ Right-click on the sequence ID for {\sl FER1\_SPIOL}.
+Select { \sl FER1\_SPIOL $\Rightarrow$ Structure $\Rightarrow$ View Structure
+$\Rightarrow$ 1A70}. A structure viewing window appears. Rotate the molecule by clicking and dragging in the structure viewing box. Zoom with the mouse scroll wheel. } \exstep{Roll the mouse cursor along the {\sl FER1\_SPIOL} sequence in the alignment. Note that if a residue in the sequence maps to one in the structure, a label will appear next to that residue in the structure viewer. Move the mouse over the structure. Placing the mouse over a part of the structure will bring up a tool tip indicating the name and number of that residue. The corresponding residue in the sequence is highlighted in black. Clicking the alpha carbon toggles the highlight and residue label on and off. Try this by clicking on a set of three or four adjacent residues so that the labels are persistent, then finding where they are in the sequence. }
+\exstep{Select {\sl Colours $\Rightarrow$ Background Colour\ldots} from the structure viewer menu and choose a suitable colour. Press OK to apply this. Select {\sl File $\Rightarrow$ Save As $\Rightarrow$ PNG} and save the image. View this with a suitable program. }
+\exstep{Select {\sl File $\Rightarrow$ View Mapping} from the structure viewer menu. A new window opens showing the residue by residue alignment between the sequence and the structure.}
+\exstep{Select {\sl File $\Rightarrow$ Save $\Rightarrow$ PDB file} and choose a new filename to save the PDB file. Once the file is saved, open the location in your file browser (or explorer window) and drag the PDB file that you just saved on to the Jalview desktop (or load it from the {\sl Jalview Desktop $\Rightarrow$ Input Alignment $\Rightarrow$ From File } menu). Verify that you can open and view the associated structure from the sequence ID pop-up menu's {\sl Structure } submenu in the new alignment window.}
+
+\exstep{Right click on the structure in the submenu and bring up the Jmol
+window.
+Explore the menu options. Try to change the style of molecular display - by first using the {\sl Jmol $\Rightarrow$ Select (n) $\Rightarrow$ All} command (where {\sl n} is the number of residues selected), and then the {\sl Jmol $\Rightarrow$ Style $\Rightarrow$ Scheme $\Rightarrow$ Ball and Stick} command.} \exstep{Use the {\sl File $\Rightarrow$ Save As .. } function to save the alignment as a Jalview Project. Now close the alignment and the structure view, and load the project file you just saved.
+
+Verify that the Jmol display is as it was when you just saved the file.}
}
-\section{Multiple Sequence Alignment}
-\label{msaservices}
-Sequences can be aligned using a range of algorithms provided by JABA web
-services. These include ClustalW\footnote{{\sl ``CLUSTAL W: improving the
-sensitivity of progressive multiple sequence alignment through sequence
-weighting, position specific gap penalties and weight matrix choice."} Thompson
-JD, Higgins DG, Gibson TJ (1994) {\sl Nucleic Acids Research} {\bf 22},
-4673-80}, Muscle\footnote{{\sl ``MUSCLE: a multiple sequence alignment method
-with reduced time and space complexity"} Edgar, R.C.
-(2004) {\sl BMC Bioinformatics} {\bf 5}, 113}, MAFFT\footnote{{\sl ``MAFFT: a
-novel method for rapid multiple sequence alignment based on fast Fourier
-transform"} Katoh, K., Misawa, K., Kuma, K. and Miyata, T. (2002) {\sl Nucleic
-Acids Research} {\bf 30}, 3059-3066. and {\sl ``MAFFT version 5:
-improvement in accuracy of multiple sequence alignment"} Katoh, K., Kuma, K.,
-Toh, H. and Miyata, T. (2005) {\sl Nucleic Acids Research} {\bf 33}, 511-518.},
-ProbCons,\footnote{PROBCONS: Probabilistic Consistency-based Multiple Sequence
-Alignment.
-Do, C.B., Mahabhashyam, M.S.P., Brudno, M., and Batzoglou, S.
-(2005) {\sl Genome Research} {\bf 15} 330-340.} T-COFFEE\footnote{T-Coffee:
-A novel method for multiple sequence alignments. (2000) Notredame, Higgins and
-Heringa {\sl JMB} {\bf 302} 205-217} and Clustal Omega.\footnote{Fast, scalable
-generation of high-quality protein multiple sequence alignments using Clustal
-Omega. Sievers F, Wilm A, Dineen DG, Gibson TJ, Karplus K, Li W, Lopez R,
-McWilliam H, Remmert M, Soding J, Thompson JD, Higgins DG (2011) {\sl Molecular
-Systems Biology} {\bf 7} 539
-\href{http://dx.doi.org/10.1038/msb.2011.75}{doi:10.1038/msb.2011.75}} Of these,
-T-COFFEE is the slowest, but also the most accurate. ClustalW is historically
-the most widely used. Muscle is faster than ClustalW and probably the most
-accurate for smaller alignments and MAFFT is probably the best for large
-alignments, however Clustal Omega, which was released in 2011, is arguably the
-fastest and most accurate tool for protein multiple alignment.
+\subsection{Superimposing Structures}
+\label{superposestructs}
+Many comparative biomolecular analysis investigations aim to determine if the
+biochemical properties of a given molecule are significantly different to its
+homologues. When structure data is available, comparing the shapes of molecules
+by superimposing them enables substructure that may impart different behaviour
+to be quickly identified. The identification of optimal 3D superpositions
+involves aligning 3D data rather than sequence symbols, but the result can
+still be represented as a sequence alignment, where columns indicate positions
+in each molecule that should be superposed to recreate the optimal 3D alignment.
+
+Jalview can employ Jmol's 3D fitting routines\footnote{See
+\href{http://chemapps.stolaf.edu/jmol/docs/?ver=12.2$\#$compare}{http://chemapps.stolaf.edu/jmol/docs/?ver=12.2$\#$compare}
+for more information.} to recreate 3D structure superpositions based on the
+correspondences defined by one or more sequence alignments involving structures shown in the Jmol display. Superposition based on the currently displayed alignment view happens automatically if a
+structure is added to an existing Jmol display using the {\sl Structure
+$\Rightarrow$ View PDB Structure $\Rightarrow$ ..}. A new Jmol view containing
+superposed structures can also be created using the {\sl Structure
+$\Rightarrow$ View all {\bf N} PDB Structures} option (when {\bf {\sl N}}
+$>$ 1) if the current selection contains two or more sequences with associated
+structures.
+\subsubsection{Obtaining the RMSD for a Superposition}
+The RMSD (Root Mean Square Deviation) is a measure of how similar the structures
+are when they are superimposed. Figure \ref{mstrucsuperposition} shows a
+superposition created during the course of Exercise \ref{superpositionex}. The
+parts of each molecule used to construct the superposition are rendered using
+the cartoon style, with other parts of the molecule drawn in wireframe. The Jmol
+console, which has been opened after the superposition was performed, shows the
+RMSD report for the superposition.
+Full information about the superposition is also outputed to the Jalview
+console.\footnote{The Jalview Java Console is opened from {\sl Tools
+$\Rightarrow$ Java Console} option in the Desktop's menu bar} This output also
+includes the precise atom pairs used to superpose structures.
-To run an alignment web service, select the appropriate method from the {\sl
-Web Service $\Rightarrow$ Alignment $\Rightarrow$ \ldots} submenu (Figure
-\ref{webservices}). For each service you may either perform an alignment with
-default settings, use one of the available presets, or customise the parameters
-with the `{\sl Edit and Run ..}' dialog box. Once the job is submitted, a
-progress window will appear giving information about the job and any errors that
-occur. After successful completion of the job, a new window is opened with the
-results, in this case an alignment. By default, the new alignment will be
-ordered in the same way as the input sequences; however, many alignment programs
-re-order the input to place homologous sequences close together. This ordering
-can be recovered using the `Original ordering' entry within the {\sl Calculate
-$\Rightarrow$ Sort } sub menu.
+\subsubsection{Choosing which part of the Alignment is used for Structural
+Superposition} Jalview uses the visible part of each alignment view to define
+which parts of each molecule are to be superimposed. Hiding a column in a view
+used for superposition will remove that correspondence from the set, and will
+exclude it from the superposition and RMSD calculation.
+This allows the selection of specific parts of the alignment to be used for
+superposition. Only columns that define a complete set of correspondences for
+all structures will be used for structural superposition, and as a consequence,
+the RMSD values generated for each pair of structures superimposed can be
+directly compared.
-\begin{figure}[htbp]
-\begin{center}
-\parbox[c]{1.5in}{\includegraphics[width=1.5in]{images/ws1.pdf}}
-\parbox[c]{2.5in}{\includegraphics[width=2.5in]{images/ws2.pdf}}
-\parbox[c]{2in}{\includegraphics[width=2in]{images/ws3.pdf}}
-\caption{{\bf Multiple alignment via web services} The appropriate method is
-selected from the menu (left), a status box appears (centre), and the results
-appear in a new window (right)}
-\label{webservices}
-\end{center}
-\end{figure}
+In order to recompute a superposition after changing a view or editing the
+alignment, select the {\sl Jmol $\Rightarrow$ Align sequences } menu option. The {\sl
+Jmol $\Rightarrow$ Superpose with ..} submenu allows you to choose which of the
+associated alignments and views are to be used to create the set of
+correspondences. This menu is useful when composing complex superpositions
+involving multi-domain and multi-chain complexes, when correspondences may be
+defined by more than one alignment.
-\subsubsection{Realignment}
+Note that these menu options appear when you have two or more structures in one Jmol viewer.
-The re-alignment option is currently only supported by ClustalW and Clustal
-Omega. When performing a re-alignment, Jalview submits the current selection to
-the alignment service complete with any existing gaps. This approach is useful
-when one wishes to align additional sequences to an existing alignment without
-any further optimisation to the existing alignment. The Re-alignment service
-provided by ClustalW in this case is effectively a simple form of profile
-alignment.
-\subsubsection{Alignments of sequences that include hidden regions}
-If the view or selected region that is submitted for alignment contains hidden
-regions, then {\bf only the visible sequences will be submitted to the service}.
-Furthermore, each contiguous segment of sequences will be aligned independently
-(resulting in a number of alignment `subjobs' appearing in the status window).
-Finally, the results of each subjob will be concatenated with the hidden regions
-in the input data prior to their display in a new window. This approach ensures
-that 1) hidden column boundaries in the input data are preserved in the
-resulting alignment - in a similar fashion to the constraint that hidden columns
-place on alignment editing (see Section \ref{lockededits}), and 2) hidden
-columns can be used to preserve existing parts of an alignment whilst the
-visible parts are locally refined.
+\begin{figure}[htbp]
+\begin{center}
+\includegraphics[width=5.5in]{images/fdxsuperposition.pdf}
+\caption{{\bf Superposition of two ferredoxin structures.} The alignment on the
+left was used by Jalview to superpose structures associated with the
+FER1\_SPIOL and FER1\_MAIZE sequences in the alignment. Parts of each structure
+used for superposition are rendered as a cartoon, the remainder rendered in
+wireframe. The RMSD between corresponding positions in the structures before and
+after the superposition is shown in the Jmol console.}
+\label{mstrucsuperposition}
+\end{center}
+\end{figure}
-\exercise{Multiple Sequence Alignment}{
-\exstep{ Close all windows and open the alignment at {\sf
-http://www.jalview.org/tutorial/unaligned.fa}. Select {\sl
-Web Service $\Rightarrow$ Alignment $\Rightarrow$ Muscle with Defaults}. A window will open giving the job status. After a short time, a second window will open with the results of the alignment.} \exstep{Select the first sequence set by clicking on the window and try running ClustalW and MAFFT (from the {\sl Web Service $\Rightarrow$ Alignment} menu) on the same initial alignment. Compare them and you should notice small differences. }
-\exstep{Select the last three sequences in the MAFFT alignment, and de-align them with {\sl Edit $\Rightarrow$ Remove All Gaps}. Press [ESC] to deselect them and then submit the view for re-alignment with ClustalW.}
-\exstep{Use [CTRL]-Z to recover the alignment of the last three sequences in the MAFFT alignment. Once the ClustalW re-alignment has completed, compare the results of re-alignment of the three sequences with their alignment in the original MAFFT result.}
-\exstep{Select columns 60 to 125 in the original MAFFT alignment and hide them. Select {\sl Web Services $\Rightarrow$ Alignment $\Rightarrow$ Mafft with Defaults} to submit the visible portion of the alignment to MAFFT. When the web service job pane appears, note that there are now two alignment job status panes shown in the window.}
-\exstep{When the MAFFT job has finished, compare the alignment of the N-terminal visible region in the result with the corresponding region of the original alignment. If you wish, select and hide a few more columns in the N-terminal region, and submit the alignment to the service again and explore the effect of local alignment on the non-homologous parts of the N-terminal region.}
-}
+\exercise{Aligning Structures using the Ferredoxin
+Sequence Alignment}{\label{superpositionex}
+\exstep{Continue with the Jalview project created in exercise
+\ref{viewingstructex}. Use the {\sl Discover PDB IDs} function to retrieve PDB
+IDs associated with the FER1\_MAIZE sequence.}
+\exstep{Once discovery has completed, use the {\sl
+View PDB Structure} submenu to view one of the PDB file associated with
+FER1\_MAIZE (eg. 3B2F)
+Jalview will give you the option of aligning the structure to the one already
+open. To superimpose the structure associated with FER1\_MAIZE with the one
+associated with FER1\_SPIOL, press the {\bf Yes} button.
-\subsection{Customising the parameters used for alignment}
+{\sl The Jmol view will update to show both structures, and one will be
+moved on to the other. If this doesn't happen, use the Align function in the Jmol submenu}}
+\exstep{Create a new view on the alignment, and hide all but columns 121
+through to 132.}
+\exstep{Use the {\sl Jmol} submenu to
+recompute the superposition using just columns 121-132 of the alignment
+(The easiest way to achieve this is to select column 121-132 and in the View
+menu selected ``All but selected region'' from the Hide options).
-JABA web services allow you to vary the parameters used when performing a
-bioinformatics analysis. For JABA alignment services, this means you are
-usually able to modify the following types of parameters:
-\begin{list}{$\bullet$}{}
-\item Amino acid or nucleotide substitution score matrix
-\item Gap opening and widening penalties
-\item Types of distance metric used to construct guide trees
-\item Number of rounds of re-alignment or alignment optimisation
-\end{list}
+{\sl Note how the molecules shift position when superposed using a short part of
+the two structures.}}
+\exstep{Compare the initial and final RMSDs for superimposing molecules with
+the small section and with the whole alignment. (The RMSD report can be
+viewed by right clicking the mouse on Jmol window, and select ``Show" and ``Measurements") Which view do you think give the best 3D
+superposition, and why ?} }
+\subsection{Colouring Structure Data Associated with Multiple Alignments and
+Views} Normally, the original view from which a particular structure view was
+opened will be the one used to colour structure data. If alignments involving
+sequences associated with structure data shown in a Jmol have multiple views, Jalview gives you full control
+over which alignment, or alignment view, is used to colour the structure
+display. Sequence-structure colouring associations are
+changed {\sl via} the {\sl View $\Rightarrow$ Colour by ..} menu, which lists all
+views associated with data shown in the embedded Jmol view. A tick is shown beside
+views currently used as colouring source, and moving the
+mouse over each view will bring it to the front of the alignment display,
+allowing you to browse available colour sources prior to selecting one. If the
+{\sl Select many views} option is selected, then multiple views can be selected as sources for colouring the
+structure data. {\sl Invert selection} and {\sl Select all views} options are also provided to quickly change between multi-view selections.
-\subsubsection{Getting help on the parameters for a service}
-Each parameter available for a method usually has a short description, which
-jalview will display as a tooltip, or as a text pane that can be opened under
-the parameter's controls. In the parameter shown in Figure
-\ref{clustalwparamdetail}, the description was opened by selecting the button on the left hand side. Online help for the
-service can also be accessed, by right clicking the button and selecting a URL
-from the pop-up menu that will open.
+Note that the {\sl Select many views} option is useful if you have different
+views that colour different areas or domains of the alignment. This option is
+further explored in Section \ref{complexstructurecolours}.
\begin{figure}[htbp]
\begin{center}
-\includegraphics[width=2.5in]{images/clustalwparamdetail.pdf}
-\caption{{\bf ClustalW parameter slider detail}. From the ClustalW {\sl Clustal $\Rightarrow$ Edit settings and run ...} dialog box. }
-\label{clustalwparamdetail}
+\includegraphics[width=5.5in]{images/mviewstructurecol.pdf}
+\caption{{\bf Choosing a different view for colouring a structure display}
+Browsing the {\sl View $\Rightarrow$ Colour by ..} menu provides full control
+of which alignment view is used to colour structures when the {\sl Colours
+$\Rightarrow$ By Sequence} option is selected.}
+\label{mviewstructurecol}
\end{center}
-\end{figure}
+\end{figure}
-\subsection{Alignment Presets}
-The different multiple alignment algorithms available from JABA vary greatly in
-the number of adjustable parameters, and it is often difficult to identify what
-are the best values for the sequences that you are trying to align. For these
-reasons, each JABA service may provide one or more presets -- which are
-pre-defined sets of parameters suited for particular types of alignment
-problem. For instance, the Muscle service provides the following presets:
-\begin{list}{$\bullet$}{}
-\item Huge
-\item Protein alignments (fastest speed)
-\item Nucleotide alignments (fastest speed)
-\end{list}
+\subsubsection{Colouring Complexes}
+\label{complexstructurecolours}
+The ability to control which multiple alignment view is used to colour
+structural data is essential when working with data relating to
+multidomain biomolecules and complexes.
-The presets are displayed in the JABA web services submenu, and can also be
-accessed from the parameter editing dialog box, which is opened by selecting
-the `{\sl Edit settings and run ...}' option from the web serviceÔøΩs menu. If you have used
-a preset, then it will be mentioned at the beginning of the job status file shown
-in the web service job progress window.
+In these situations, each chain identified in the structure may have a different
+evolutionary history, and a complete picture of functional variation can
+only be gained by integrating data from different alignments on the same
+structure view. An example of this is shown in Figure
+\ref{mviewalcomplex}, based on data from Song et. al.\footnote{Structure of
+DNMT1-DNA Complex Reveals a Role for Autoinhibition in Maintenance DNA Methylation. Jikui Song, Olga Rechkoblit, Timothy H. Bestor, and Dinshaw J. Patel.
+{\sl Science} 2011 {\bf 331} 1036-1040
+\href{http://www.sciencemag.org/content/331/6020/1036}{DOI:10.1126/science.1195380}}
-\subsubsection{Alignment Service Limits}
-Multiple alignment is a computationally intensive calculation. Some JABA server
-services and service presets only allow a certain number of sequences to be
-aligned. The precise number will depend on the server that you are using to
-perform the alignment. Should you try to submit more sequences than a service
-can handle, then an error message will be shown informing you of the maximum
-number allowed by the server.
+\begin{figure}[htbp]
+\begin{center}
+\includegraphics[]{images/mchainstructureview.pdf}
+\caption{{\bf The biological assembly of Mouse DNA Methyltransferase-1 coloured
+by Pfam alignments for its major domains} Alignments for each domain within the
+Uniprot sequence DNMT1\_MOUSE have been used to visualise sequence conservation
+in each component of this protein-DNA complex. Instructions for recreating this figure are given in exercise \ref{dnmtcomplexex}. }
+\label{mviewalcomplex}
+\end{center}
+\end{figure}
-\subsection{User defined Presets}
-Jalview allows you to create your own presets for a particular service. To do
-this, select the `{\sl Edit settings and run ...}' option for your service,
-which will open a parameter editing dialog box like the one shown in Figure
-\ref{jwsparamsdialog}.
+\exercise{Colouring a Protein Complex to Explore Domain-Domain
+Interfaces}{\label{dnmtcomplexex}
-The top row of this dialog allows you to browse the existing presets, and
-when editing a parameter set, allows you to change its nickname. As you
-adjust settings, buttons will appear at the top of the parameters dialog that
-allow you to Revert or Update the currently selected user preset with your changes, Delete the current preset, or Create a new preset, if none exists with the given name. In addition to the parameter set name, you can also provide a short
-description for the parameter set, which will be shown in the tooltip for the
-parameter set's entry in the web services menu.
+\exstep{Download the PDB file at
+\textsf{\url{http://www.jalview.org/tutorial/DNMT1\_MOUSE.pdb}} to your desktop.
-\begin{figure}[htbc]
-\center{
-\includegraphics[width=3in]{images/jvaliwsparamsbox.pdf}
-\caption{{\bf Jalview's JABA alignment service parameter editing dialog box}.}
-\label{jwsparamsdialog} }
-\end{figure}
+This is the biological unit for PDB ID 3pt6, as identified by the PDBe's PISA
+server.}
-\subsubsection{Saving parameter sets}
-When creating a custom parameter set, you will be asked for a file name to save
-it. The location of the file is recorded in the Jalview user preferences in the
-same way as a custom alignment colourscheme, so when Jalview is launched again,
-it will show your custom preset amongst the options available for running the
-JABA service.
+\exstep{Launch the Jalview desktop and ensure you have at least 256MB of
+free memory available.
-%
-% \exercise{Creating and using user defined presets}{\label{createandusepreseex}
-% \exstep{Import the file at
-% \textsf{http://www.jalview.org/tutorial/fdx\_unaligned.fa} into jalview.}
-% \exstep{Use the `{\slDiscover Database Ids}' function to recover the PDB cross
-% references for the sequences.}
-% \exstep{Align the sequences using the default ClustalW parameters.}
-% \exstep{Use the `{\sl Edit and run..}'
-% option to open the ClustalW parameters dialog box, and create a new preset using
-% the following settings:
-% \begin{list}{$\bullet$}{}
-% \item BLOSUM matrix (unchanged)
-% \item Gap Opening and End Gap penalties = 0.05
-% \item Gap Extension and Separation penalties = 0.05
-% \end{list}
-%
-% As you edit the parameters, buttons will appear on the dialog box
-% allowing you revert your changes or save your settings as a new parameter
-% set.
-%
-% Before you save your settings, remeber to give them a meaningful name by editing
-% the text box at the top of the dialog box.
-% }
-% \exstep{Repeat the alignment using your new parameter set by selecting it from
-% the {\sl ClustalW Presets menu}.}
-% \exstep{These sequences have PDB structures associated with them, so it is
-% possible to compare the quality of the alignments.
-%
-% Use the {\sl View all {\bf N}
-% structures} option to calculate the superposition of 1fdn on 1fxd for both
-% alignments (refer to section \ref{superposestructs} for instructions). Which
-% alignment gives the best RMSD ? }
-% \exstep{Apply the same alignment parameter settings to the example alignment
-% (available from \textsf{http://www.jalview.org/examples/uniref50.fa}).
-%
-% Are there differences ? If not, why not ?
-% }
-% }
+{\sl Use the following webstart link:
-\section{Protein alignment conservation analysis}
-\label{aacons}
-The {\sl Web Service $\Rightarrow$ Conservation} menu controls the computation
-of up to 17 different amino acid conservation measures for the current alignment
-view. The JABAWS AACon Alignment Conservation Calculation Service, which is used
-to calculate these scores, provides a variety of standard measures described by
-Valdar in 2002\footnote{Scoring residue conservation. Valdar (2002) {\sl
-Proteins: Structure, Function, and Genetics} {\bf 43} 227-241.} as well as an efficient implementation of the SMERFs
-score developed by Manning et al. in 2008.\footnote{SMERFS Score Manning et al. {\sl BMC
-Bioinformatics} 2008, {\bf 9} 51 \href{http://dx.doi.org/10.1186/1471-2105-9-51}{doi:10.1186/1471-2105-9-51}}
+\href{http://www.jalview.org/services/launchApp?jvm-max-heap=1G}{http://www.jalview.org/services/launchApp?jvm-max-heap=1G}.}
-\subsubsection{Enabling and disabling AACon calculations}
-When the AACon Calculation entry in the {\sl Web Services $\Rightarrow$
-Conservation} menu is ticked, AACon calculations will be performed every time
-the alignment is modified. Selecting the menu item will enable or disable
-automatic recalculation.
+{\sl Alternatively in the Development section of the Jalview web site
+(\href{http://www.jalview.org/development/development-builds}{http://www.jalview.org/development/development-builds})
+in the ``latest official build'' row in the table, go to the
+``Webstart'' column, click on ``G2''.}}
+\exstep{Retrieve the following {\bf full} PFAM alignments: PF02008, PF01426
+(make sure you select the {\sl PFAM {\bf (Full)}} source). These will each be retrieved into their own alignment window.} \exstep{Drag the URL or file of the structure you downloaded in
+step 1 onto one of the alignments to associate it with the mouse sequence in
+that Pfam domain family.}
+\exstep{For every DNMT1\_MOUSE sequence in the alignment, use the sequence
+ID popup menu's {\sl Structure} submenu to view the DNMT1\_MOUSE structure for the associated mouse sequence. When given the option, {\bf view all of the structures in the same Jmol viewer}. Check the contents of the {\sl View $\Rightarrow$ Colour by ..} submenu to see what alignments can be used to
+colour the sequence.}
+\exstep{Repeat the previous two steps for each of
+the other alignments. In each case, when performing the `View DNMT1\_MOUSE.pdb'
+step, Jalview will ask if you wish to create a new Jmol view. You should
+respond `No', {\bf ensuring that each sequence fragment is associated with the same Jmol view}.}
+\exstep{Pick a different colourscheme for each alignment, and use the {\sl
+Colour by ..} submenu to ensure they are all used to colour the complex shown
+in the Jmol window.}
+\exstep{The final step needed to reproduce the shading in Figure
+\ref{mviewalcomplex} is to use the {\sl Colour $\Rightarrow$ By
+Annotation } option in each alignment window to shade the alignment by the
+{\bf Conservation} annotation row. This function was described in section
+\ref{colourbyannotation}.
-\subsubsection{Configuring which AACon calculations are performed}
-The {\sl Web Services $\Rightarrow$ Conservation $\Rightarrow$ Change AACon
-Settings ...} menu entry will open a web services parameter dialog for the
-currently configured AACon server. Standard presets are provided for quick and
-more expensive conservation calculations, and parameters are also provided to
-change the way that SMERFS calculations are performed.
-AACon settings for an alignment are saved in Jalview projects along with the
-latest calculation results.
+Ensure that you first disable the {\sl View $\Rightarrow$ Show Features} menu option, or you may not see any colour changes in the associated structure.
-\subsubsection{Changing the server used for AACon calculations}
-If you are working with alignments too large to analyse with the public JABAWS
-server, then you will most likely have already configured additional JABAWS
-servers. By default, Jalview will chose the first AACon service available from
-the list of JABAWS servers available. If available, you can switch to use
-another AACon service by selecting it from the {\sl Web Services $\Rightarrow$
-Conservation $\Rightarrow$ Switch Server} submenu.
+{\sl Note: Choose a different shading scheme for each
+alignment so that the regions of strong physicochemical conservation are highlighted. This
+kind of shading will reveal conserved regions of interaction between domains
+in the structure.}}
+\exstep{Save your work as a Jalview project and verify that it can be opened again by starting another Jalview Desktop instance, and dragging the saved project into the desktop window.}
+
+% {\sl Note: This exercise relies on new features introduced in Jalview 2.7. If
+% you notice any strange behaviour when trying out this exercise, it may be a
+% bug (see
+% \href{http://issues.jalview.org/browse/JAL-1008}{http://issues.jalview.org/browse/JAL-1008}
+% for one relating to highlighting of positions in the alignment window).}
+}
% TODO
+\chapter{Protein Prediction Analysis}
+\label{proteinprediction}
\section{Protein Secondary Structure Prediction}
\label{protsspredservices}
Protein secondary structure prediction is performed using the
be mapped back onto the visible parts of the sequence, to ensure a single frame
of reference is maintained in your analysis.
+\section{Protein Disorder Prediction}
+\label{protdisorderpred}
+
+Disordered regions in proteins were classically thought to correspond to
+``linkers'' between distinct protein domains, but disorder can also play a role in
+function. The {\sl Web Services $\Rightarrow$ Disorder} menu in the alignment window
+allows access to protein disorder prediction services provided by the configured
+JABAWS servers.
+
+\subsection{Disorder Prediction Results}
+Each service operates on sequences in the alignment to identify regions likely
+to be unstructured or flexible, or alternately, fold to form globular domains.
+As a consequence, disorder predictor results include both sequence features and
+sequence associated alignment annotation rows. Section \ref{featannot} describes
+the manipulation and display of these data in detail, and Figure
+\ref{alignmentdisorder} demonstrates how sequence feature shading and
+thresholding (described in Section \ref{featureschemes}) can be used to
+highlight differences in disorder prediction across aligned sequences.
+
\exercise{Secondary Structure Prediction}{
\label{secstrpredex}
\exstep{ Open the alignment at \url{http://www.jalview.org/tutorial/alignment.fa}. Select the sequence {\sl FER\_MESCR} by clicking on the sequence ID. Then select {\sl Web Services $\Rightarrow$ Secondary Structure Prediction $\Rightarrow$ JNet Secondary Structure Prediction} from the alignment window menu. A status window will appear and after some time (about 2-4 min) a new window with the JPred prediction will appear. Note that the number of sequences in the results window is many more than in the original alignment as JNet performs a PSI-BLAST search to expand the prediction dataset.
{\sl Note: you may want to keep this data for use in exercise \ref{viewannotfileex}.}
}
-}
-
-\section{Protein Disorder Prediction}
-\label{protdisorderpred}
-
-Disordered regions in proteins were classically thought to correspond to
-'linkers' between distinct protein domains, but disorder can also play a role in
-function. The {\sl Web Services $\Rightarrow$ Disorder} menu in the alignment window
-allows access to protein disorder prediction services provided by the configured
-JABAWS servers.
-
-\subsection{Disorder prediction results}
-Each service operates on sequences in the alignment to identify regions likely
-to be unstructured or flexible, or alternately, fold to form globular domains.
-As a consequence, disorder predictor results include both sequence features and
-sequence associated alignment annotation rows. Section \ref{featannot} describes
-the manipulation and display of these data in detail, and {\bf Figure
-\ref{alignmentdisorder}} demonstrates how sequence feature shading and
-thresholding (described in Section \ref{featureschemes}) can be used to
-highlight differences in disorder prediction across aligned sequences.
+\exstep{
+In the original alignment that you loaded in step 1, {\bf select all} sequences,
+then open the {\bf Sequence ID $\Rightarrow$ Selection } submenu and select the
+{\bf Add Reference Annotation} option.
+
+The JNet predictions for the sequences should now be visible in the original
+alignment.} }
\begin{figure}[htbp]
\begin{center}
\end{center}
\end{figure}
-\subsubsection{Navigating large sets of disorder predictions}
+\subsubsection{Navigating Large Sets of Disorder Predictions}
-{\bf Figure \ref{alignmentdisorderannot}} shows a single sequence annotated with
+Figure \ref{alignmentdisorderannot} shows a single sequence annotated with
a range of disorder predictions. Disorder prediction annotation rows are
associated with a sequence in the same way as secondary structure prediction
results. When browsing an alignment containing large numbers of disorder
\begin{figure}[htbp]
\begin{center}
\includegraphics[width=5in]{images/disorderpredannot.pdf}
-\caption{{\bf Annotation rows for several disorder predictions on a sequence}. A zoomed out view of a prediction for a single sequence. The sequence is shaded to highlight disorderd regions (brown and grey), and the line plots below the Sequence show the raw scores for various disorder predictors. Horizontal lines on each graph mark the level at which disorder predictions become significant. }
+\caption{{\bf Annotation rows for several disorder predictions on a sequence}. A
+zoomed out view of a prediction for a single sequence. The sequence is shaded to highlight disordered regions (brown and grey), and the line plots below the Sequence show the raw scores for various disorder predictors. Horizontal lines on each graph mark the level at which disorder predictions become significant. }
\label{alignmentdisorderannot}
\end{center}
\end{figure}
-\subsection{Disorder predictors provided by JABAWS 2.0}
+\subsection{Disorder Predictors provided by JABAWS 2.0}
For full details of each predictor and the results that Jalview can display,
please consult
\href{http://www.jalview.org/help/html/webServices/proteinDisorder.html}{Jalview's
\textbf{Globular Domain} features mark long stretches of order. \textbf{Dydx} annotation rows gives the first order derivative of smoothed score. Values above 0 indicates
residue is disordered.
-\textbf{Smoothed Score and Raw Score} annotation rows give the smoothed and raw scores used to create the differential signal that
-indicates the presence of unstructured regions. These are hidden
-by default, but can be shown by right-clicking on the alignment
-annotation panel and selecting \textbf{Show hidden annotation}.
-
-\section{Features and Annotation}
-\label{featannot}
-Features and annotations are additional information that is overlaid on the sequences and the alignment. Generally speaking, annotations are associated with columns in the alignment. Features are associated with specific residues in the sequence.
-
-Annotations are shown below the alignment in the annotation panel, and often reflect properties of the alignment as a whole. The Conservation, Consensus and Quality scores are examples of dynamic annotation, so as the alignment changes, they change along with it. Conversely, sequence features are properties of the individual sequences, so they do not change with the alignment, but are shown mapped on to specific residues within the alignment.
-
-Features and annotation can be interactively created, or retrieved from external
-data sources. DAS (the Distributed Annotation System) is the primary source of
-sequence features, whilst webservices like JNet (see \ref{jpred} above) can be used to analyse a given sequence or alignment and generate annotation for it.
-
-
-\subsection{Creating sequence features}
-Sequence features can be created simply by selecting the area in a sequence (or sequences) to form the feature and selecting {\sl Selection $\Rightarrow$ Create Sequence Feature } from the right-click context menu (Figure \ref{features}). A dialogue box allows the user to customise the feature with respect to name, group, and colour. The feature is then associated with the sequence. Moving the mouse over a residue associated with a feature brings up a tool tip listing all features associated with the residue.
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=2in]{images/feature1.pdf}
-\includegraphics[width=2.5in]{images/feature2.pdf}
-\includegraphics[width=1.5in]{images/feature3.pdf}
-\caption{{\bf Creating sequence features.} Features can readily be created from selections via the context menu and are then displayed on the sequence. }
-\label{features}
-\end{center}
-\end{figure}
-
-Creation of features from a selection spanning multiple sequences results in the creation of one feature per sequence. Each feature remains associated with it's own sequence.
-
-\subsection{Customising feature display}
-
-Feature display can be toggled on or off by selecting the {\sl View
-$\Rightarrow$ Show Sequence Features} menu option. When multiple features are
-present it is usually necessary to customise the display. Jalview allows the
-display, colour, rendering order and transparency of features to be modified
-{\sl via} the {\sl View $\Rightarrow$ Feature Settings\ldots} menu option. This
-brings up a dialogue window (Figure \ref{custfeat}) which allows the
-visibility of individual feature types to be selected, colours changed (by
-clicking on the colour of each sequence feature type) and the rendering order
-modified by dragging feature types to a new position in the list. Dragging the
-slider alters the transparency of the feature rendering. The Feature
-Settings dialog also includes functions for more advanced feature shading
-schemes and buttons for sorting the alignment according to the distribution of
-features. These capabilities are described further in sections
-\ref{featureschemes} and \ref{featureordering}.
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=4in]{images/features4.pdf}
-\caption{{\bf Multiple sequence features.} An alignment with JPred secondary structure prediction annotation below it, and many sequence features overlaid onto the aligned sequences. The tooltip lists the features annotating the residue below the mouse-pointer.}
-\end{center}
-\end{figure}
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=4in]{images/features5.pdf}
-\caption{{\bf Customising sequence features.} Features can be recoloured, switched on or off and have the rendering order changed. }
-\label{custfeat}
-\end{center}
-\end{figure}
-
-\subsection{Sequence Feature File Formats}
-
-Jalview supports the widely used GFF tab delimited format\footnote{see
-http://www.sanger.ac.uk/resources/software/gff/spec.html} and its own Jalview
-Features file format for the import of sequence annotation. Features and
-alignment annotation are also extracted from other formats such as Stockholm,
-and AMSA. URL links may also be attached to features. See the online
-documentation for more details of the additional capabilities of the jalview
-features file.
-
-\exercise{Creating features}{
-\exstep{Open the alignment at \textsf{http://www.jalview.org/tutorial/alignment.fa}. We know that the Cysteine residues at columns 97, 102, 105 and 135 are involved in iron binding so we will create them as features. Navigate to column 97, sequence 1. Select the entire column by clicking in the ruler bar. Then right-click on the selection to bring up the context menu and select {\sl Selection $\Rightarrow$ Create Sequence Feature}. A dialogue box will appear.
-}
-\exstep{
-Enter a suitable Sequence Feature Name (e.g. ``Iron binding site") in the
-appropriate box. Click on the Feature Colour bar to change the colour if
-desired, add a short description (``One of four Iron binding Cysteines") and press OK. The features will then appear on the sequences. } \exstep{Roll the mouse cursor over the new features. Note that the position given in the tool tip is the residue number, not the column number. To demonstrate that there is one feature per sequence, clear all selections by pressing [ESC] then insert a gap in sequence 3 at column 95. Roll the mouse over the features and you will see that the feature has moved with the sequence. Delete the gap you created.
-}
-\exstep{
-Add a similar feature to column 102. When the feature dialogue box appears, clicking the Sequence Feature Name box brings up a list of previously described features. Using the same Sequence Feature Name allows the features to be grouped.}
-\exstep{Select {\sl View $\Rightarrow$ Feature Settings\ldots} from the
-alignment window menu. The Sequence Feature Settings window will appear. Move
-this so that you can see the features you have just created. Click the check
-box for ``Iron binding site" under {\sl Display} and note that display of this
-feature type is now turned off. Click it again and note that the features are
-now displayed. Close the sequence feature settings box by clicking OK or
-Cancel.} }
-
-\subsection{Creating user defined annotation}
-
-Annotations are properties that apply to the alignment as a whole and are visualized on rows in the annotation panel.
-To create a new annotation row, right click on the annotation label panel and select the {\sl Add New Row} menu option (Figure \ref{newannotrow}). A dialogue box appears. Enter the label to use for this row and a new row will appear.
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=1.3in]{images/annots1.pdf}
-\includegraphics[width=2in]{images/annots2.pdf}
-\caption{{\bf Creating a new annotation row.} Annotation rows can be reordered by dragging them to the desired place.}
-\label{newannotrow}
-\end{center}
-\end{figure}
-
-To create a new annotation, first select all the positions to be annotated on the appropriate row. Right-clicking on this selection brings up the context menu which allows the insertion of graphics for secondary structure ({\sl Helix} or {\sl Sheet}), text {\sl Label} and the colour in which to present the annotation (Figure \ref{newannot}). On selecting {\sl Label} a dialogue box will appear, requesting the text to place at that position. After the text is entered, the selection can be removed and the annotation becomes clearly visible\footnote{When annotating a block of positions, the text can be partly obscured by the selection highlight. Pressing the [ESC] key clears the selection and the label is then visible.}. Annotations can be coloured or deleted as desired.
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=2in]{images/annots3.pdf}
-\includegraphics[width=2in]{images/annots4.pdf}
-\includegraphics[width=2in]{images/annots5.pdf}
-\caption{{\bf Creating a new annotation.} Annotations are created from a selection on the annotation row and can be coloured as desired.}
-\label{newannot}
-\end{center}
-\end{figure}
-
-\exercise{Annotating alignments}{
-\exstep{Load the alignment at \textsf{http://www.jalview.org/tutorial/alignment.fa}. Right-click on the annotation label for {\sl Conservation} to bring up the context menu and select {\sl Add New Row}. A dialogue box will appear asking for {\sl Label for annotation}. Enter ``Iron binding site" and click OK. A new, empty, row appears.
-}
-\exstep{
-Navigate to column 97. Select column 97 on the new annotation row. Right click on the selection and select {\sl Label} from the context menu. Enter ``Fe" in the box and click OK. Right-click on the selection again and select {\sl Colour}. Choose a colour from the colour chooser dialogue and click OK. Press [ESC] to remove the selection.
-}
-\exstep{ Select columns 70-77 on the annotation row. Right-click and choose {\sl Sheet} from the context menu. You will be prompted for a label. Enter ``B" and press OK. A new line showing the sheet as an arrow appears. The colour of the label can be changed but not the colour of the sheet arrow.
-}
-\exstep{Right click on the annotation row that you just created. Select {\sl Export Annotation} and, in the {\bf Export Annotation} dialog box that will open, select the Jalview format and click the [To Textbox] button.
-
-The format for this file is given in the Jalview help. Press [F1] to open it, and find the ``Annotations File Format'' entry in the ``Alignment Annotations'' section of the contents pane. }
-
-\exstep{Export the file to a text editor and edit the file to change the name of the annotation row. Save the file and drag it onto the alignment view.}
-\exstep{Try to add an additional helix somewhere along the row by editing the file and re-importing it.
-{\sl Hint: Use the {\bf Export Annotation} function to view what helix annotation looks like in a jalview annotation file.}}
-\exstep{Use the {\sl Alignment Window $\Rightarrow$ File $\Rightarrow$ Export Annotations...} function to export all the alignment's annotation to a file.}
-\exstep{Open the exported annotation in a text editor, and use the {\bf Annotation File Format} documentation to modify the style of the Conservation, Consensus and Quality annotation rows so they appear as several lines on a single line graph.
-{\sl Hint: You need to change the style of annotation row in the first field of the annotation row entry in the file, and create an annotation row grouping to overlay the three quantitative annotation rows.}
-}
-\label{viewannotfileex}\exstep{Recover or recreate the secondary structure
-prediction that you made in exercise \ref{secstrpredex}. Use the {\sl File $\Rightarrow$ Export Annotation} function to view the Jnet secondary structure prediction annotation row. Note the {\bf SEQUENCE\_REF} statements surrounding the row specifying the sequence association for the annotation. } }
-
-\section{Importing features from databases}
-\label{featuresfromdb}
-Jalview supports feature retrieval from public databases either directly or {\sl
-via} the Distributed Annotation System (DAS\footnote{http://www.biodas.org/}).
-It includes built in parsers for Uniprot and EMBL records retrieved from the
-EBI. Sequences retrieved from these sources using the sequence fetcher (see
-Section \ref{fetchseq}) will already possess features.
-
-\subsection{Sequence Database Reference Retrieval}
-\label{fetchdbrefs}
-Jalview maintains a list of external database references for each sequence in
-an alignment. These are listed in a tooltip when the mouse is moved over the
-sequence ID when the {\sl View $\Rightarrow$ Sequence ID Tooltip $\Rightarrow$
-Show Database Refs } option is enabled. Sequences retrieved using the sequence
-fetcher will always have at least one database reference, but alignments
-imported from an alignment file generally have no database references.
-
-\subsubsection{Database References and Sequence Coordinate Systems}
-
-Jalview displays features in the local sequence's coordinate system which is
-given by its `start' and `end'. Any sequence features on the sequence will be
-rendered relative to the sequence's start position. If the start/end positions
-do not match the coordinate system from which the features were defined, then
-the features will be displayed incorrectly.
-
-\subsubsection{Viewing and exporting a sequence's database annotation}
-
-You can export all the database cross references and annotation terms shown in
-the sequence ID tooltip for a sequence by right-clicking and selecting the {\sl
-[Sequence ID] $\Rightarrow$ Sequence details \ldots} option from the popup menu.
-A similar option is provided in the {\sl Selection} sub-menu allowing you to
-obtain annotation for the sequences currently selected.
-
-\parbox[l]{3.4in}{
-The {\sl Sequence Details
-\ldots} option will open a window containing the same text as would be shown in
-the tooltip window, including any web links associated with the sequence. The
-text is HTML, and options on the window allow the raw code to be copied and
-pasted into a web page.}
-\parbox[c]{3in}{
-\centerline{\includegraphics[width=2.2in]{images/seqdetailsreport.pdf}}}
-
-\subsubsection{Automatically discovering a sequence's database references}
-Jalview includes a function to automatically verify and update each sequence's
-start and end numbering against any of the sequence databases that the {\sl
-Sequence Fetcher} has access to. This function is accessed from the {\sl
-Webservice $\Rightarrow$ Fetch DB References} sub-menu in the Alignment
-window. This menu allows you to query either the set of {\sl Standard
-Databases}, which includes EMBL, Uniprot, the PDB, and the currently selected
-DAS sequence sources, or just a specific datasource from one of the submenus.
-When one of the entries from this menu is selected, Jalview will use the ID
-string from each sequence in the alignment or in the currently selected set to
-retrieve records from the external source. Any sequences that are retrieved are
-matched against the local sequence, and if the local sequence is found to be a
-sub-sequence of the retrieved sequence then the local sequence's start/end
-numbering is updated. A new database reference mapping is created, mapping the
-local sequence to the external database, and the local sequence inherits any
-additional annotation retrieved from the database sequence.
-
-The database retrieval process terminates when a valid mapping is found for a
-sequence, or if all database queries failed to retrieve a matching sequence.
-Termination is indicated by the disappearance of the moving progress indicator
-on the alignment window. A dialog box may be shown once it completes which
-lists sequences for which records were found, but the sequence retrieved from
-the database did not exactly contain the sequence given in the alignment (the
-{\sl ``Sequence not 100\% match'' dialog box}).
-
-\exercise{Retrieving Database References}{
-\exstep{Load the example alignment at http://www.jalview.org/tutorial/alignment.fa}
-\exstep{Verify that there are no database references for the sequences by first
-checking that the {\sl View $\Rightarrow$ Sequence ID Tooltip $\Rightarrow$ Show
-Database IDs} option is selected, and then mousing over each sequence's ID.}
-\exstep{Use the {\sl Webservice $\Rightarrow$ Fetch DB References} menu option to retrieve database IDs for the sequences.}
-\exstep{Examine the tooltips for each sequence in the alignment as the retrieval progresses - note the appearance of new database references.}
-\exstep{Once the process has finished, save the alignment as a Jalview Project.}
-\exstep{Now close all the windows and open the project again, and verify that the database references and sequence features are still present on the alignment}
-
-\exstep{View the {\sl Sequence details \ldots} report for the FER1\_SPIOL sequence and for the whole alignment. Which sequences have web links associated with them ?}
-
-}
-
-\subsection{Retrieving Features {\sl via} DAS}
-\label{dasfretrieval}
-Jalview includes a client to retrieve features from DAS annotation servers. To
-retrieve features, select {\sl View $\Rightarrow$ Feature Settings\ldots} from the alignment window menu. Select the {\sl DAS Settings} tab in the Sequence Feature Settings Window (Figure \ref{das}). A list of DAS sources compiled from the currently configured DAS registry\footnote{By default, this will be the major public DAS server registry maintained by the Sanger Institute: http://www.dasregistry.org} is shown in the left hand pane. Highlighting an entry on the left brings up information about that source in the right hand panel.
-
-\begin{figure}[htbp]
-\begin{center}
-\includegraphics[width=2.5in]{images/das1.pdf}
-\includegraphics[width=2.5in]{images/das2.pdf}
-\caption{{\bf Retrieving DAS annotations.} DAS features are retrieved using the {\sl DAS Settings} tab (left) and their display customised using the {\sl Feature Settings} tab (right).}
-\label{das}
-\end{center}
-\end{figure}
-
-Select appropriate DAS sources as required then click on {\sl Fetch DAS
-Features}. If you know of additional sources not listed in the configured
-registry, then you may add them with the {\sl Add Local Source} button. Use
-the {\sl Authority},{\sl Type}, and {\sl Label} filters to restrict the list
-of sources to just those that will return features for the sequences in the
-alignment.
-
-Following DAS feature retrieval, the {\sl Feature Settings} panel takes on a
-slightly different appearance (Figure \ref{das} (right)). Each data source is
-listed and groups of features from one data source can be selected/deselected
-by checking the labeled box at the top of the panel.
-
-
-\subsubsection{The Fetch Uniprot IDs dialog box}
-\label{discoveruniprotids}
-If any sources are selected which refer to Uniprot coordinates as their reference system, then you may be asked if you wish to retrieve Uniprot IDs for your sequence. Pressing OK instructs Jalview to verify the sequences against Uniprot records retrieved using the sequence's ID string. This operates in much the same way as the {\sl Web Service $\Rightarrow$ Fetch Database References } function described in Section \ref{fetchdbrefs}. If a sequence is verified, then the start/end numbering will be adjusted to match the Uniprot record to ensure that features retrieved from the DAS source are rendered at the correct position.
-
-\subsubsection{Rate of feature retrieval}
-Feature retrieval can take some time if a large number of sources is selected and if the alignment contains a large number of sequences. This is because Jalview only queries a particular DAS source with one sequence at a time, to avoid overloading it. As features are retrieved, they are immediately added to the current alignment view. The retrieved features are shown on the sequence and can be customised as described previously.
-
-
-\exercise{Retrieving features with DAS}{
-\label{dasfeatretrexcercise}
-\exstep{Load the alignment at
-\textsf{http://www.jalview.org/tutorial/alignment.fa}. Select {\sl View
-$\Rightarrow$ Feature Settings \ldots} from the alignment window menu. Select
-the {\sl DAS Settings} tab. A long list of available DAS sources is listed. Select a small number, eg Uniprot, DSSP, signalP and netnglyc. Click OK. A window may prompt whether you wish Jalview to map the sequence IDs onto Uniprot IDs. Click {\sl Yes}. Jalview will start retrieving features. As features become available they will be mapped onto the alignment. } \exstep{If Jalview is taking too long to retrieve features, the process can be cancelled with the {\sl Cancel Fetch} button. Rolling the mouse cursor over the sequences reveals a large number of features annotated in the tool tip. Close the Sequence Feature Settings window. }
-\exstep{Move the mouse over the sequence ID panel. Non-positional features such as literature references and protein localisation predictions are given in the tooltip, below any database cross references associated with the sequence.}
-\exstep{Search through the alignment to find a feature with a link symbol next to it. Right click to bring up the alignment view popup menu, and find a corresponding entry in the {\sl Link } sub menu. }
-% TODO this doesn't work ! \includegraphics[width=.3in]{images/link.pdf}
-
-\exstep{
-Select {\sl View $\Rightarrow$ Feature Settings\ldots} to reopen the Feature Settings window. All the loaded feature types should now be displayed. Those at the top of the list are drawn on top of those below, obscuring them in the alignment view where they overlap. Move the feature settings window so that the alignment is visible and uncheck some of the feature types by clicking the tick box in the display column. Observe how the alignment display changes. Note that unselected feature types do not appear in the tool tip.
-}
-\exstep{Reorder the features by dragging feature types up and down the order in the Feature Settings panel. e.g. Click on {\sl CHAIN} then move the mouse downwards to drag it below {\sl DOMAIN}. Note that {\sl DOMAIN} is now shown on top of {\sl CHAIN} in the alignment window. Drag {\sl METAL} to the top of the list. Observe how the cysteine residues are now highlighted as they have a {\sl METAL} feature associated with them.
-}
-
-\exstep{Press the {\sl Optimise Order} button. The features will be ordered according to increasing length, placing features that annotate shorter regions of sequence higher on the display stack.}
-
-\exstep{Select {\sl File $\Rightarrow$ Export Features\ldots} from the Alignment window. You can choose to export the retrieved features as a GFF file, or Jalview's own Features format.
-% TODO: describe working with features files and GFF
-}
-}
-
-\subsection{Colouring features by score or description
-text}
-\label{featureschemes}
-Sometimes, you may need to visualize the differences in information carried by
-sequence features of the same type. This is most often the case when features
-of a particular type are the result of a specific type of database query or calculation. Here, they may also carry information within their textual description, or most commonly for calculations, a score related to the property being investigated. Jalview can shade sequence
-features using a graduated colourscheme in order to highlight these variations.
-In order to apply a graduated scheme to a feature type, select the `Graduated
-colour' entry in the Sequence {\sl Feature Type}'s popup menu, which is opened by
-right-clicking the {\sl Feature Type} or {\sl Color} in the {\sl Sequence Feature Settings} dialog box. Two types
-of colouring styles are currently supported: the default is quantitative
-colouring, which shades each feature based on its score, with the highest
-scores receiving the `Max' colour, and the lowest scoring features coloured
-with the `Min' colour. Alternately, you can select the `Colour by label'
-option to create feature colours according to the description text associated
-with each feature. This is useful for general feature types - such as
-Uniprot's `DOMAIN' feature - where the actual type of domain is given in the
-feature's description.
-
-Graduated feature colourschemes can also be used to exclude low or
-high-scoring features from the alignment display. This is done by choosing your
-desired threshold type (either above or below), using the drop-down menu in the
-dialog box. Then, adjust the slider or enter a value in the text box to set the
-threshold for displaying this type of feature.
+\textbf{Smoothed Score and Raw Score} annotation rows give the smoothed and raw scores used to create the differential signal that
+indicates the presence of unstructured regions. These are hidden
+by default, but can be shown by right-clicking on the alignment
+annotation panel and selecting \textbf{Show hidden annotation}.
-The feature settings dialog box allows you to toggle between a graduated and
-simple feature colourscheme using the pop-up menu for the feature type. When a
-graduated scheme is applied, it will be indicated in the colour column for
-that feature type - with coloured blocks or text to indicate the colouring
-style and a greater than ($>$) or less than ($<$) symbol to indicate when a
-threshold has been defined.
+\exercise{Protein Disorder Prediction}{
+%\label{protdispredex}
-\subsection{Using features to re-order the alignment}
-\label{featureordering}
-The presence of sequence features on certain sequences or in a particular
-region of an alignment can quantitatively identify important trends in
-the aligned sequences. In this case, it is more useful to
-re-order the alignment based on the number of features or their associated scores, rather than simply re-colour the aligned sequences. The sequence feature settings
-dialog box provides two buttons: `Seq sort by Density' and `Seq sort by
-Score', that allow you to reorder the alignment according to the number of
-sequence features present on each sequence, and also according to any scores
-associated with a feature. Each of these buttons uses the currently displayed
-features to determine the ordering, but
-if you wish to re-order the alignment using a single type of feature, then you can do this from the {\sl Feature Type}'s
-popup menu. Simply right-click the type's style in the Sequence Feature Settings dialog
-box, and select one of the {\sl Sort by Score} and {\sl Sort by Density}
-options to re-order the alignment. Finally, if a specific region is selected,
-then only features found in that region of the alignment will be used to
-create the new alignment ordering.
+\exstep{Open the alignment at
+\url{http://www.jalview.org/tutorial/interleukin7.fa}. } also available at
-\exercise{Shading and sorting alignments using sequence features}{
-\label{shadingorderingfeatsex}
-\exstep{Re-load the alignment from \ref{dasfeatretrexcercise}.
-}
-\exstep{Open the
-feature settings panel, and, after first clearing the current
-selection, press the {\em Seq Sort by Density} button a few times.}
-\exstep{Use the DAS fetcher to retrieve the Kyte and Doolittle Hydrophobicity
-scores for the protein sequences in the alignment.
-{\sl Hint: the nickname for the das source is `KD$\_$hydrophobicity'.}}
-\exstep{Change the feature settings so only the hydrophobicity features are
-displayed. Mouse over the annotation and also export and examine the GFF and
-Jalview features file to better understand how the hydrophobicity measurements
-are recorded.}
-\exstep{Apply a {\sl Graduated Colour} to the hydrophobicity annotation to
-reveal the variation in average hydrophobicity across the alignment.}
-\exstep{Select a range of alignment columns, and use one of the sort by feature buttons to order the alignment according to that region's average
-hydrophobicity.}
-\exstep{Save the alignment as a project, for use in exercise
-\ref{threshgradfeaturesex}.} }
-
-\exercise{Shading alignments with combinations of graduated feature
-colourschemes}{
-\label{threshgradfeaturesex}
-\exstep{Reusing the annotated alignment from exercise
-\ref{shadingorderingfeatsex}, experiment with the colourscheme threshold to
-highlight the most, or least hydrophobic regions. Note how the {\sl Colour} icon for the {\sl Feature Type} changes when you change the threshold type and press OK.}
-\exstep{Change the colourscheme so
-that features at the threshold are always coloured grey, and the most
-hydrophobic residues are coloured red, regardless of the threshold value
-({\em hint - there is a switch on the dialog to do this for you}).}
-\exstep{Enable the Uniprot {\em chain} annotation in the feature settings
-display and re-order the features so it is visible under the hydrophobicity
-annotation.}
-\exstep{Apply a {\sl Graduated Colour} to the {\em chain}
-annotation so that it distinguishes the different canonical names associated
-with the mature polypeptide chains.}
-\exstep{Export the alignment's sequence features using the Jalview and GFF file formats, to see how the different types of graduated feature
-colour styles are encoded. }
-}
+
+\url{http://www.jalview.org/tutorial/training-materials/2014/Dundee/Oct/interleukin7.fa}
+
+\exstep{Run the DisEMBL disorder predictor {\slvia} the {\slWeb Services
+$\Rightarrow$ Disorder Prediction } submenu.}
+
+\exstep{Use {\sl Sequence ID $\Rightarrow$ Structure $\Rightarrow$ Discover PDB
+IDs} to retrieve all the PDB structures for the sequences.}
+
+\exstep{Open and align
+the structures for all sequences.
+
+{\sl Hint: see \ref{viewAllStructures} to see how to do this.}}
+
+\exstep{Compare the disorder predictions to the structure data by mapping any
+available temperature factors to the alignment {\sl via} the {\sl Sequence ID
+Popup $\Rightarrow$ Selection $\Rightarrow$ Add reference annotation} option.}
+
+\exstep{Apply the IUPred disorder prediction method}
+\exstep{Use the {\sl Per
+sequence option} in the {\sl Colour $\Rightarrow$ By annotation \ldots} dialog to shade
+the sequences by the long and short disorder predictors.
+Do the two methods agree with the structure ?}}
+
+\chapter{DNA and RNA Sequences}
+\label{dnarna}
\section{Working with DNA}
\label{workingwithnuc}
Jalview was originally developed for the analysis of protein sequences, but
sequence fetcher (see Section \ref{fetchseq}) are also parsed in order to
identify codon regions and extract peptide products. Furthermore, Jalview
records mappings between protein sequences that are derived from regions of a
-nucleotide sequence. Mappings are used to to transfer annotation between
+nucleotide sequence. Mappings are used to transfer annotation between
nucleic acid and protein sequences, and to dynamically highlight regions in
one sequence that correspond to the position of the mouse pointer in another.
%TODO Working with Nucleic acid sequences and structures.
Jalview has limited knowledge of the capabilities of the programs that
are made available to it {\sl via} web services, so it is up to you, the user,
to decide which service to use when working with nucleic acid sequences. The
-table below shows which alignment programs are most appropriate
+table shows which alignment programs are most appropriate
for nucleotide alignment. Generally, all will work, but some may be more suited
to your purposes than others. We also note that none of these include
support for taking RNA secondary structure prediction into account when aligning
-sequences. We expect that in the future, Jalview will fully support secondary
-structure aware RNA alignment.
-
+sequences (but will be providing services for this in the future!)
\begin{table}{}
\centering
\begin{tabular}{|l|c|l|}
}
-\subsection{Coding regions from EMBL records}
+\subsection{Coding Regions from EMBL Records}
Many EMBL records that can be retrieved with the sequence fetcher contain exons.
Coding regions will be marked as features on the EMBL nucleotide sequence, and
\subsubsection{Retrieval of Protein or DNA Cross References}
The {\sl Calculate $\Rightarrow$ Get Cross References } function is only available when Jalview recognises that there are protein/DNA cross-references present on sequences in the alignment. When selected, it retrieves the cross references from the alignment's dataset (a set of sequence and annotation metadata shared between alignments) or using the sequence database fetcher. This function can be used for EMBL sequences containing coding regions to open the Uniprot protein products in a new alignment window. The new alignment window that is opened to show the protein products will also allow dynamic highlighting of codon positions in the EMBL record for each residue in the protein product(s).
-\subsubsection{Retrieval of protein DAS features on coding regions}
+\subsubsection{Retrieval of Protein DAS Features on Coding Regions}
The Uniprot cross-references derived from EMBL records can be used by Jalview to visualize protein sequence features directly on nucleotide alignments. This is because the database cross references include the sequence coordinate mapping information to correspond regions on the protein sequence with that of the nucleotide contig. Jalview will use the Uniprot accession numbers associated with the sequence to retrieve features, and then map them onto the nucleotide sequence's coordinate system using the coding region location.
\end{center}
\end{figure}
-\exercise{Visualizing protein features on coding regions}
+\exercise{Visualizing Protein Features on Coding Regions}
{
-\exstep{Use the sequence fetcher to retrieve EMBL record V00488.}
-\exstep{Ensure that {\sl View $\Rightarrow$ Show Sequence Features} is checked and change the alignment view format to Wrapped mode so the distinct exons can be seen.}
-\exstep{Open the {\sl DAS Settings} tab in the {\sl Sequence Feature Settings\ldots} window and fetch features for V00488 from the Uniprot reference server, and any additional servers that work with the Uniprot coordinate system.}
+\exstep{Use the sequence fetcher to retrieve EMBL record D49489.}
+\exstep{Ensure that {\sl View $\Rightarrow$ Show Sequence Features} is checked and change the
+alignment view format to Wrapped mode so the distinct exons can be seen.}
+\exstep{Open the {\sl DAS Settings} tab in the {\sl Sequence Feature Settings\ldots}
+window {\sl View $\Rightarrow$ Features setting}
+and fetch features for D49489 from the Uniprot reference server,
+and any additional servers that work with the Uniprot coordinate system.}
\exstep{Mouse over the features retrieved, note that they have been mapped onto the coding regions, and in some cases broken into several parts to cover the distinct exons.}
-\exstep{Open a new alignment view containing the Uniprot protein product with {\sl Calculate $\Rightarrow$ Get Cross References $\Rightarrow$ Uniprot } and examine the database references and sequence features. Experiment with the interactive highlighting of codon position for each residue.
-}
+\exstep{Open a new alignment view containing the Uniprot protein product
+with {\sl Calculate $\Rightarrow$ Get Cross References $\Rightarrow$ Uniprot }
+and examine the database references and sequence features. Experiment with the
+interactive highlighting of codon position for each residue.}
}
% \section{Working with RNA}
% \label{workingwithrna}
% \subsection{Using local PDB files}
% \section{Pairwise alignments}
- \end{document}
+\section{Working with RNA}
+Jalview allows the creation of RNA secondary structure annotation, and includes
+the VARNA secondary structure viewer for the display of RNA base pair diagrams.
+It also allows the extraction of RNA secondary structure from 3D data when
+available.
+
+\subsection{Performing RNA Secondary Structure Predictions}
+Secondary structure consensus calculations can be performed by enabling the
+VIENNA service {\sl via} the {\sl Web Services $\Rightarrow$ Secondary
+Structure} menu. These consensus structures are created by analysing the
+covariation patterns in all visible sequences on the alignment. For more
+information see the VIENNA documentation.
+
+\begin{figure}[htbp]
+\begin{center}
+\label{rnaviennaservice}
+\includegraphics[width=5in]{images/rnaViennaServiceWindow.pdf}
+
+\caption{Secondary structure consensus calculations can be performed by enabling the
+VIENNA service {\sl via} the {\sl Web Services $\Rightarrow$ Secondary
+Structure} menu.}
+
+\end{center}
+\end{figure}
+
+\begin{figure}[htbp]
+\begin{center}
+\label{rnaviennaaltpairs}
+\includegraphics[width=5in]{images/rnaViennaAlternateProbs.pdf}
+
+\caption{VIENNA can calculate alternate RNA base pairing probabilities. These
+are shown in Jalview as tool-tips on the RNA secondary structure probability
+score.}
+
+\end{center}
+\end{figure}
+
+
+\exercise{Viewing RNA Structures}
+{ \label{viewingrnaex}
+
+\exstep{Import RF00162 from Rfam (Full) using Fetch sequence(s) option in File
+menu.}
+\exstep{Select {\sl Colour by RNA Helices} to shade the alignment by
+the secondary structure annotation provided by Rfam.}
+
+\exstep{Open VARNA with {\sl Structure $\Rightarrow$ View Structure
+$\Rightarrow$ RNA Secondary Structure}.
+In the VARNA Structures Manager toggle between (i) secondary structure
+(alignment) (with gaps) and (ii) trimmed secondary structure (alignment).
+Explore the difference between trimmed and untrimmed views.
+Click on different residues and located them in the sequence alignment window.}
+
+\exstep{In the VARNA Structures Manager, right click on display window to
+bring up the pop up context menu. Investigate option within File, Export, Display
+and Edit sections.}
+
+\exstep{Perform a secondary structure prediction in {\sl Web Services}. Enable
+the VIENNA consensus calculation via the {\sl Web Services} menu and select
+Change RNAAIiFold settings option.}
+% Compare this with the annotationline provided by Rfam.
+
+\exstep{Edit the VIENNA calculation settings to show
+Base Pair probabilities. Explore how editing the alignment affects the consensus
+calculation.}
+
+\exstep{Import 2GIS from PDB database using Fetch sequence(s) option.}
+\exstep{Click on a sequence in Sequence ID panel and select {\sl Structure
+$\Rightarrow$ View Structure $\Rightarrow$ 2GIS}, to view the structure in Jmol
+window. Click on different residues and located them in the sequence alignment window.}
+%\exstep{Add and link a Jmol structure view
+%for Bacillus\_amyloliquef.9 for 3NPB (from the PDB). Display the secondary
+%structure along-side the consensus structure for the alignment by adding
+%reference annotation from the 3D structure.
+
+%{\sl Hint: You need to make sure the RNAview service is enabled in your {\sl
+%Structure} preferences to obtain RNA secondary structure annotation from PDB
+%files.}}
+
+ }
+
+\chapter{Webservices}
+\label{jvwebservices}
+The term ``Webservices'' refers to a variety of data exchange
+mechanisms based on HTTP.\footnote{HTTP: Hyper-Text Transfer Protocol.}
+
+\parbox[c]{4.5in}{Jalview can exploit public webservices to access databases
+remotely, and also submit data to public services by opening pages with your web browser. These types of
+services are `one-way', {\sl i.e.} data is either sent to the webservice or
+retrieved from it by Jalview. The desktop application can also interact
+with `two-way' remote analysis services in order to offload computationally
+intensive tasks to High Performance Computing facilities. Most of these two-way
+services are provided by {\bf Ja}va {\bf B}ioinformatics {\bf A}nalysis {\bf
+W}eb {\bf S}ervice (JABAWS) servers\footnote{See
+http://www.compbio.dundee.ac.uk/jabaws for more information and to download
+your own server.}, which provides an easily installable system for performing
+a range of bioinformatics analysis tasks. }
+\parbox[c]{1.75in}{\includegraphics[width=1.65in]{images/wsmenu.pdf}}
+
+\subsection{One-Way Web Services}
+
+There are two types of one way service in Jalview. Database services,
+which were introduced in in Section \ref{fetchseq}, provide sequence and
+alignment data. They can also be used to add sequence IDs to an alignment
+imported from a local file, prior to further annotation retrieval, as described
+in Section \ref{featuresfromdb}. A second type of one way service is provided
+by Jalview's DAS sequence feature retrieval system, which is described
+in Section \ref{dasfretrieval}.
+% The final type of one way service are sequence
+% and ID submission services.
+% exemplified by the `Envision2 Services' provided
+% by the ENFIN Consortium\footnote{ENFIN is the European Network for Functional
+% INtegration. Please see http://www.enfin.org for more information. }.
+
+% \subsubsection{One-way submission services}
+% Jalview can use the system's web browser to submit sets of sequences and
+% sequence IDs to web based applications. Single sequence IDs can be passed to
+% a web site using the user definable URL links listed under the {\sl
+% Links} submenu of the sequence ID popup menu. These are configured
+% in the {\sl Connections} tab of the {\sl Preferences} dialog box.
+%
+% The Envision 2 services presented in the webservice menu provides are the first
+% example of one-way services where multiple sequences or sequence IDs can be
+% sent. The {\sl Web service $\Rightarrow$ Envision 2 Services} menu entry
+% provides two sub-menus that enable you to submit the sequences or IDs
+% associated with the alignment or just the currently selected sequences to one
+% of the Envision2 workflows. Selecting any one will open a new browser window on
+% the Envision2 web application. The menu entries and their tooltips provide
+% details of the Envision2 workflow and the dataset set that will be submitted
+% ({\sl i.e.} the database reference type, or associated sequence subset). Please
+% note, due to technical limitations, Jalview can currently only submit small
+% numbers of sequences to the workflows - if no sequence or ID submissions are
+% presented in the submenus, then try to select a smaller number of sequences to
+% submit.
+
+\subsection{Remote Analysis Web Services}
+Remote analysis services enable Jalview to use external computational
+facilities. There are currently three types of service - multiple sequence
+alignment, protein secondary structure prediction, and alignment analysis.
+Many of these are provided by JABA servers, which are described at the end of
+this section. In all cases, Jalview will construct a job based on the alignment
+or currently selected sequences, ask the remote server to run the job, monitor
+status of the job and, finally, retrieve the results of the job and display
+them. The Jalview user is kept informed of the progress of the job through a
+status window.
+
+Currently, web service jobs and their status windows are not stored in Jalview
+Project Files\footnote{This may be rectified in future versions.}, so it is
+important that you do not close Jalview whilst a job is running. It is also
+essential that you have a continuous network connection in order to
+successfully use web services from Jalview, since it periodically checks the
+progress of running jobs.
+
+
+\subsection{JABA Web Services for Sequence Alignment and Analysis}
+\label{jabaservices}
+JABA stands for ``JAva Bioinformatics Analysis'', which is a system developed
+by Peter Troshin and Geoff Barton at the University of Dundee for running
+computationally intensive bioinformatics analysis programs. A JABA installation
+typically provides a range of JABA web services (JABAWS) for use by other
+programs, such as Jalview.
+
+Exercises in the remainder of this section will demonstrate the simplest way of
+installing JABA on your computer, and configuring Jalview so it can access the JABA services. If you
+need any further help or more information about the services, please go to the
+\href{http://www.compbio.dundee.ac.uk/jabaws}{JABAWS home page}.
+%% \subsubsection{Aims}
+%% \begin{list}{$\bullet$}{}
+%% \item Gain experience using the different alignment services provided by
+% JABA
+%%\item Learn about the way that Jalview stores user presets for JABA services
+%%\item Learn how to install JABA services and configure Jalview to access them
+%%\end{list}
+
+\subsection{Changing the Web Services Menu Layout}
+\label{changewsmenulayout}
+If you are working with a lot of different JABA services, you may wish to change
+the way Jalview lays out the web services menu. You can do this from the Web
+Services tab of the {\sl Preferences} dialog box.
+
+\exercise{Changing the Layout of the Web Services Menu}{
+\label{changewsmenulayoutex}
+\exstep{Make sure you have loaded an alignment into Jalview, and examine the
+current layout of the alignment window's {\sl Web Service} menu.}
+\exstep{Open the preferences dialog box and select the web services tab.}
+\exstep{Ensure the {\sl Enable JABAWS services} checkbox is selected, and unselect
+the {\sl Enable Enfin Services} checkboxes.}
+\exstep{Hit {\sl Refresh Services} to update the web services menu -- once the
+progress bar has completed, open the {\sl Web Service} menu to view the changes.}
+\exstep{Select the {\sl Index by host} checkbox and refresh the services once again.
+
+{\sl Observe the way the layout of the JABAWS Alignment submenu changes.}
+}
+\exstep{Do the same with the {\sl Index by type} checkbox.}
+}
+
+Jalview provides these options for configuring the layout of the {\sl Web Service}
+menu because different Jalview users may have access to a different number of
+JABA services, and each will have their own preference regarding the layout of
+the menu.
+
+\begin{figure}[htbc]
+\begin{center}
+\includegraphics[width=3in]{images/jvjabawsconfig.pdf}
+\caption{{\bf The Jalview Web Services preferences panel.} Options are provided
+for configuring the list of JABA servers that Jalview will use, enabling and
+disabling Enfin services, and configuring the layout of the web services
+menu.}
+\label{jvjabawsconfig}
+\end{center}
+\end{figure}
+
+
+\subsubsection{Testing JABA services}
+The JABAWS configuration dialog shown in Figure \ref{jvjabawsconfig} has colour
+codes to indicate whether the Desktop can access the server, and whether all
+services advertised by the server are functional. The colour codes are:
+
+\begin{list}{$\bullet$}{}
+ \item Red - Server cannot be contacted or reports a connection error.
+ \item Amber - Jalview can connect, but one or more services are non-functional.
+ \item Green - Server is functioning normally.
+\end{list}
+ %TODO green and a tick, red and a cros, amber and a value indicating whether its all going
+
+Test results from JABAWS are reported on Jalview's console output (opened from
+the Tools menu). Tests are re-run every time Jalview starts, and when the
+[Refresh Services] button is pressed on the Jalview JABAWS configuration panel.
+
+\subsubsection{Resetting the JABA Services Setting to their Defaults}
+Once you have configured a JABAWS server and selected the OK button of the
+preferences menu, the settings will be stored in your Jalview preferences file,
+along with any preferences regarding the layout of the web services menu. If
+you should ever need to reset the JABAWS server list to its defaults, use the
+`Reset Services' button on the Web Services preferences panel.
+
+\subsection{Running your own JABA Server}
+You can download and run JABA on your own machine using the `VMWare' or
+VirtualBox virtual machine environments. If you would like to learn how to do
+this, there are full instructions at the
+\href{http://www.compbio.dundee.ac.uk/jabaws/}{JABA web site}.
+
+\exercise{Installing a JABA Virtual Machine on your Computer}{
+\label{jabawsvmex}{\sl This tutorial will demonstrate the simplest way of
+installing JABA on your computer, and configuring Jalview so it can access the JABA services.
+
+{\bf Prerequisites}
+
+You will need a copy of VMWare Player/Workstation/Fusion on your machine.
+}
+
+\exstep{If you do not have VMWare player installed, download it from
+www.vmware.com (this takes a few minutes -- you will need to register and wait
+for an email with a download link).}
+\exstep{Download the JABA virtual appliance archive called `jaba-vm.zip' from
+\textsf{http://www.compbio.dundee.ac.uk/jabaws/archive/jabaws-vm.zip}
+
+WARNING: This is large (about 300MB) and will take some time to download.
+}
+\exstep{Unpack the archive's contents to a place on your machine with at least
+2GB of free space.
+
+(On Windows, right click on the archive, and use the 'Extract archive..' option).
+}
+\exstep{Open the newly extracted directory and double click the VMWare virtual
+machine configuration file (jabaws.vcf). This will launch the VMWare player.
+}
+\exstep{Once VMWare player has started up, it may ask the question ``Did you move or copy
+this virtual appliance?'' -- select `Copy'.}
+\exstep{You may be prompted to download the VMWare linux tools. These are not
+necessary, so close the window or click on `Later'.}
+\exstep{You may also be prompted to install support for one or more devices (USB
+or otherwise). Say `No' to these options.}
+\exstep{Once the machine has loaded, it will display a series of IP addresses
+for the different services provided by the VM. Make a note of the JABAWS URL --
+this will begin with `http:' and end with `/jabaws''.}
+}
+
+\exercise{Configuring Jalview to Access your new JABAWS Virtual Appliance}{
+\label{confnewjabawsappl}
+\exstep{Start Jalview (If you have not done so already).}
+\exstep{Enable the Jalview Java Console by selecting its option from the Tools
+menu.
+
+{\sl Alternately, use the System Java console if you have
+configured it to open when Jalview is launched, {\sl via} your system's Java
+preferences (under the `Advanced' tab on Windows).}}
+\exstep{Open the {\sl Preferences} dialog and locate the Web Services tab.}
+\exstep{Add the URL for the new JABAWS server you started in Exercise
+\ref{jabawsvmex} to the list of JABAWS urls using the `New Service
+URL' button.}
+\exstep{You will be asked if you want to test the service. Hit `Yes' to do this
+-- you should then see some output in the console window.
+
+{\sl Take a close look at the output in the console. What do you think is
+happening?}
+}
+\exstep{Hit OK to save your preferences -- you have now added a new JABA
+service to Jalview!}
+\exstep{Try out your new JABA services by loading the ferredoxin sequences from
+http://www.jalview.org/tutorial/alignment.fa}
+\exstep{Launch an alignment using one
+of the JABA methods provided by your server. It will be listed under the JABAWS Alignment submenu of the {\sl Web Service} menu on the alignment window.
+
+{\sl Note: You can watch the JABA VM appliance's process working by opening the
+process monitor on your system. (On Windows XP, this involves right-clicking the
+system clock and opening the task manager -- then selecting the 'Processes' tab
+and sort by CPU).}
+}
+}
+
+\end{document}
git://source.jalview.org / jalview-manual.git / blobdiff