+\parbox[c]{4in}{
+PNG is Portable Network Graphics. This output option produces an image that can be easily included in web pages and incorporated in presentations using e.g. Powerpoint or Open Office. It is a bitmap image so does not scale and is unsuitable for use on posters, or in publications.
+
+For submission of alignment figures to journals, please use EPS\footnote{If the journal complains, {\em insist}.}.
+}
+\parbox[c]{4in}{\centerline{\includegraphics[width=3in]{images/image_png.pdf}}
+\par \centerline{Zoom Detail of PNG image.}}
+
+ \exercise{Graphical Output}{
+ \label{graphicex}
+\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 when zoomed.
+(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.}
+\exstep{Experiment with Jalview's other output options: try exporting an alignment view as 'BioJS', which employs the BioJS Multiple Sequence Alignment viewer. When would you use this type of export option ?}
+\exstep{Working with embedded BioJSON data. Drag and drop (or load via the file browser) the 'BioJS' HTML file in the previous step. Compare the original and imported alignment views - are there differences ?}
+\bf See the video at:
+\url{http://www.jalview.org/training/Training-Videos}.
+}
+
+% left out for Glasgow 2016
+% \newpage
+%
+% \section{Summary - the rest of the manual}
+%
+% The first few chapters have covered the basics of Jalview operation: from
+% starting the program, importing, exporting, selecting, editing and colouring
+% aligments, to the generation of figures for publication, presentation and web
+% pages.
+%
+% The remaining chapters in the manual cover:
+%
+% \begin{list}{$\circ$}{}
+% \item{Chapter \ref{featannot} covers the creation, manipulation and visualisation
+% of sequence and alignment annotation, and retrieval of sequence and feature data
+% from databases.}
+% \item {Chapter \ref{msaservices} explores the range of multiple alignment
+% programs offered via Jalview's web services, and introduces the use of
+% AACon for protein multiple alignment conservation analysis.}
+% \item {Chapter \ref{alignanalysis} introduces Jalview's built in tools for
+% multiple sequence alignment analysis, including trees, PCA, and alignment
+% conservation analysis. }
+% \item {Chapter \ref{3Dstructure} demonstrates the structure visualization
+% capabilities of Jalview.}
+% \item {Chapter \ref{proteinprediction} introduces protein sequence based
+% secondary structure and disorder prediction tools, including JPred.}
+% \item {Chapter \ref{dnarna} covers the special functions and
+% visualization techniques for working with RNA alignments and protein coding
+% sequences.}
+% \item {Chapter \ref{jvwebservices} provides instructions on the
+% installation of your own Jalview web services.}
+% \end{list}
+
+\chapter{Annotation and Features}
+\label{featannot}
+Annotations and features are additional information that is
+overlaid on the sequences and the alignment.
+Generally speaking, annotations reflect properties of the alignment as a
+whole, often associated
+with columns in the alignment. Features are often associated with specific
+residues in the sequence.
+
+Annotations are shown below the alignment in the annotation panel, the
+properties are often based on the alignment.
+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. Webservices like JPred (see \ref{jpred} above) can be used to
+analyse a given sequence or alignment and generate annotation for it.
+
+
+\section{Conservation, Quality, Consensus and other Annotation}
+\label{annotationintro}
+Jalview automatically calculates several quantitative alignment annotations
+which are displayed as histograms below the multiple sequence alignment columns.
+Conservation, quality and consensus 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.
+
+\subsubsection{Conservation Annotation}
+
+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 '+'.
+
+\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{Occupancy Annotation}
+
+Alignment occupancy simply reflects the number of residues aligned at each column
+in the multiple sequence alignment. To see this annotation you may first need to enable it by ticking the {\sl Occupancy} check-box
+in the {\sl Visual} tab in Jalview's {\sl Preferences} before opening an alignment. Occupancy is particularly useful in conjunction with
+the {\sl Select/Hide by Annotations} dialog since it allows the view to be filtered to exclude regions of the alignment with a high proportion of gaps.
+
+\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}
+
+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 dialog 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 dialog 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=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}
+
+\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}
+
+\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 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.
+
+
+\exercise{Annotating Alignments}{
+ \label{annotatingalignex}
+\exstep{Load the alignment at \textsf{http://www.jalview.org/tutorial/alignment.fa}.
+Right-click on the label name of the {\sl Conservation} annotation row to
+bring up the context menu and select {\sl Add New Row}. A dialog box will
+appear asking for {\sl Annotation Name} and {\sl Annotation Description}.
+Enter ``Iron binding site" and click {\sl 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 {\sl OK}. Right-click on the selection again
+and select {\sl Colour}.
+Choose a colour from the colour chooser dialog
+and click {\sl OK}. Press [ESC] to remove the selection.
+
+{\sl Note: depending on your Annotation sort settings, your newly
+created annotation row might 'jump' to the top or bottom of the annotation
+panel. Just scroll up or down to find it again - the column you marked will
+still be selected. }
+
+}
+\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 {\sl 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 in the annotation row that you just
+created.
+Select {\sl Export Annotation} in context menu and, in the Export Annotation
+dialog box that will open, select the Jalview format and click the {\sl [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{Open a text editor and copy the annotation text into the editor.
+Edit the text by changing the name of the annotation row and save the file.}
+\exstep{Drag the file onto the alignment in Jalview and check the annotation
+panel.} \exstep{Return to the text editor, add an additional helix somewhere
+along the row, save the file and re-importing it into Jalview as previously.
+{\sl Hint: Use the 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. Save the file.}
+\exstep{Open the exported annotation in a text editor, and use the 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. Create an annotation row grouping to overlay the
+three quantitative annotation rows.}
+}
+\exstep{{\bf Homework once you have completed exercise
+\ref{secstrpredex}:}
+\label{viewannotfileex}
+
+Recover or recreate the secondary structure predictions that you made from
+JPred. Use the {\sl File $\Rightarrow$ Export Annotation} function to view the Jnet secondary structure prediction annotation row.
+
+Note: the
+SEQUENCE\_REF statements surrounding the row specifying the sequence association for the
+annotation.
+}
+\bf See the video at:
+\url{http://www.jalview.org/training/Training-Videos}.}
+
+
+\section{Sequence Features}
+Sequence features are annotation associated with a specific sequence - often marking a specific region, such as a domain or binding site. Jalview allows features to 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 dialog 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 its own sequence.
+
+
+
+\section{Importing Features from Databases}
+\label{featuresfromdb}
+Jalview supports feature retrieval from public databases.
+It includes built in parsers for Uniprot, Ensembl 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.
+
+
+\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}
+
+
+\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.}