Jalview Help

• Menus
  • File
  • Edit
  • Font
  • View
  • Colour
  • Calculate
  • Align
   
• Command line parameters
 
• Applet parameters
 
• Input/Output

• Selecting/editing operations
   
• Changing the appearance

• Colour schemes
   
• Fonts
   
• Calculations
   
• Multiple alignments
   
•  SRS access
   
• The Alignment Window
 
• The status bar
 
• The ID panel

 

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Menus

File menu

• Input from the command line
• Input from a URL
• Input from a local file (application only)
• Save to a local file (application only)
• Save postscript to a local file (application only)
• Input via a text box (cut and paste)
• Alignment output in a text box (cut and paste)
• Alignment output by email
• Postscript output by email
• File formats supported
• Applet parameters
 

Input from the command line

When run as an application jalview takes the name of an alignment file on the command line.  The format is :
 

    java jalview.AlignFrame <alignfile> File|URL <format> [-mail <mailserver>

                  -srsserver <srsserver> -database <srsdatabase>]

For people using the script file jalview.bat or Jalview the syntax is
 

    Jalview <alignfile> <format>

<alignfile>  is the name of the alignment file which has to be in one of the supported formats.  The allowed formats are MSF, FASTA, PIR, CLUSTAL and BLC and are described in more detail here.

File|URL  If  you are reading from a local file use the File switch here.  If you are reading a URL use the URL switch.

<format>  This must be one of MSF, FASTA, PIR CLUSTAL, BLC, PFAM, MSP

Examples:

     1) For a URL:

           java jalview.AlignFrame http://circinus.ebi.ac.uk:6543/jalview/llym.msf URL MSF

   2) For a local file

           Jalview 1lym.blc BLC

           java jalview.AlignFrame 1lym.pir File PIR

<mailserver>  This defaults to circinus.ebi.ac.uk.  For far away places and behind firewalls insert your own mail server.

<srsserver>  The default is the EBI srs server :

            srs.ebi.ac.uk:5000/srs5bin/cgi-bin/   .

If you use your own srs server then this option takes the location (minus the http://) of the wgetz program.  In most cases this will be

            your.hostname/srs5bin/cgi-bin/

<database>  The default database is swall which is probably EBI specific.  Change this to your relevant database.

See here for more details about SRS access.

Example for access to the sanger SRS site and the pfam database :

        java jalview.AlignFrame myfile.pfam File PFAM -srsserver www.sanger.ac.uk/srs5bin/cgi-bin/ -database pfamseq

Input from a URL

See Input from the command line for details of how to do this.

 

Input from a local file (application only)

Selecting this option brings up a window where you can type in your local file and select the right format.  Pressing the 'Apply' button loads up the alignment in a new window.  Input formats allowed are MSF, FASTA, PIR, BLC, CLUSTAL, PFAM and MSP.  Further details about formats are here.




Output to a local file (application only)

This option allows you to save your alignment as text to a local file using much the same procedure as reading in a file.  WARNING:  files can be overwritten with no prompting.



Outputting postscript to a local file (application only)

You can save your coloured alignment as postscript using this option. A window appears where you can select the font and fontsize you wish to use as well as whether the output orientation is portrait or landscape.



Input via a text box

As applets can't write or read local files I have provided a way of inputting alignments by cutting and pasting.
Select your alignment from your local text editor or xterm and paste directly into jalview.  You then have to select the format your alignment text is in and click apply to tell jalview to interpret the output.  If the format is correct a new alignment window will appear.

Unix selection:  'cat myalignfile' will display the alignment file on screen.  Select the alignment text with the mouse.  Move the mouse over the text input window and press the middle button.  You should now have transferred your alignment to the jalview input box.

Windows selection:  Open up your alignment in a text editor (notepad, wordpad or whatever).  Select all the text with the mouse and type CTRL-C to copy it.  Move your mouse over the jalview text input window and type CTRL-V.  The alignment text should now be transferred to the input window.

 

Alignment Output by Email

Again, as applets can't write local files jalview provides output via email.    Selecting the 'Mail alignment...' option from the File menu brings up a window where you can enter your email address and select the output format you want.  Pressing the 'Apply' button initiates the mail transfer and will send you a text version of your alignment.



Alignment output via a text box

Similarly to the textbox input option the text version of your alignment can be output via a java text box.  See the 'Input via a text box' description for how to cut and paste your alignments.



Postscript Output by Email

A coloured version of your alignment can be obtained by email.  Selecting the 'Mail postscript...' option in the file menu brings up a window where you can enter your email address.  You can then select your choice of fonts and the orientation of the paper.   Pressing the Apply button generates the postscript and mails it to your account.  For large alignments this can take quite a while and can generate large postscript files (>1Mb).

 

File formats supported

The formats supported are
 

• MSF (GCG output with no checksum),
• CLUSTAL (Clustalw output),
• FASTA (common and simple format),
• PIR (less common but almost as simple format) ,
• BLC (AMPS output)
• PFAM (simple and has the advantage of including start-end points).

 For more details look here.

 

Applet parameters

If you are running jalview via the button applet provided in the distribution then there are a number of parameters you can set to define different sequence groups and colour schemes.  These are described here:

The html to do this is:

<APPLET codebase="http://srs.ebi.ac.uk/~michele/jalview/"

     code="jalview.ButtonAlignApplet"

     width = 1000

     height = 35>

  <param name=input value="http://srs.ebi.ac.uk/~michele/jalview/lipase.msf">

  <param name=type value="URL">

  <param name=format value="MSF">

</APPLET>

 

Description of the html parameters

The <APPLET> tag

<APPLET codebase="http://www.ebi.ac.uk/~michele/jalview/"

        code="jalview.ButtonAlignApplet"

        width = 150

        height = 40>

This line defines where the code for the applet lies (www.ebi.ac.uk) and what java class is to be called
(jalview.ButtonAlignApplet).

The width and height parameters define how many pixels the applet should take up in the browser.  In this case they  should be enough to display a button.

This CODEBASE method loads classes as and when it needs them so is probably suited for slow net connections.  An
alternative is to load all the classes at once in a jar file (~100kb).  This makes the applet slow to start but subsequent
operations faster.
 
 

To use a jar file:
 

<APPLET ARCHIVE="http://www.ebi.ac.uk/~michele/jalview/jalview.jar"

        CODE="jalview.ButtonAlignApplet"

        WIDTH= 250

        HEIGHT = 40>

 

Specifying an alignment file and format

 <param name=input value="http://www.ebi.ac.uk/jalview/test.msf">

 <param name=type value="URL">

 <param name=format value="MSF">

These parameters describe where to find the alignment and what format it is in.

    input - The URL of the alignment to be viewed
    type   -  Whether the alignment is a file or a URL.  Use URL here as File is not allowed for security reasons.
    format - the format of the alignment.  Allowable values are MSF, CLUSTAL, FASTA and PIR.
 
 

Advanced html parameters

The applet above reads in a protein alignment in fasta format that also contains some secondary structure predictions and definitions. The alignment is split into two groups which are displayed differently.

The protein alignment has the residue text displayed and is coloured according to each residues agreement with the consensus sequence. The secondary structure predictions/definitions have no text displayed but are coloured according to the structural element (helix/sheet).

The consensus calculation is done over a subset of the sequences in the alignment (the protein sequences). This is also specified in the html. The html to do this is :
 

<APPLET archive="http://srs.ebi.ac.uk:5000/~michele/jalview/jalview.jar"

        code="jalview.ButtonAlignApplet" width = 100 height = 35>

<PARAM name=input value="http://srs.ebi.ac.uk:5000/~michele/jalview/test2.fa">

<PARAM name=type value="URL"><PARAM name=format value="FASTA">

<PARAM name="groups" value="2"><PARAM name="group1" value="1-36:PID:true:true:false">

<PARAM name="group2" value="37-45:SECONDARY:false:true:true">

<PARAM name=fontsize value="10">

<PARAM name="mailServer" value = "srs.ebi.ac.uk">

<PARAM name="Consensus" value="1-36">

<PARAM name="srsServer" value = "srs.ebi.ac.uk:5000/srs5bin/cgi-bin/">

<PARAM name="database" value = "swall">

</APPLET>

 

Defining groups

<param name="groups" value="2">

<param name="group1" value="1-36:PID:true:true:false">

<param name="group2" value="37-45:SECONDARY:true:false:false">

    groups -  the number of groups defined
    groupn  - the group definition for group n.  If there are N groups defined with the groups parameter then there must be
N groupn parameter lines.<br>
    The value string for the groupn parameter is split into 5 fields separated by colons.  The fields are:

        <sequences in group> : <colour scheme> : <display boxes> : <display text> :  <colour text>
 
 

Allocating sequences to a group

The first field defines which sequences are to be in this group.   A few examples are the most informative here I think.

       20-26                    Sequences 20,21,22,23,2425,26 are in the group
       1,2,4                       Sequences 1,2 and 4 are in the group
       10-15,2,6-7         Sequences 2,6,7,10,11,12,13,14,15 are in the group
 
 

Defining a group colour scheme

This colour scheme will apply to the residue boxes or the residue text depending on whether those parameters are set

Allowed values are

    ZAPPO                  - colour residues by physio-chemical properties
    PID                         - colour residues by percentage identity agreement with consensus sequence
    BLOSUM62          - colour residues by agreement with consensus or +ve BLOSUM62 score with consensus residue
    HYDROPHOBIC  - colour residues by hydrophobicity (red - hydrophobic, blue - hydrophilic).
    SECONDARY      -  colour residues by secondary structure (H - magenta, E - yellow).
 
 

Displaying residue boxes

The third parameter can be true or false according to whether you want residue boxes displayed or not

Displaying text

The 4th parameter can be true or false according to whether you want the residue text displayed or not.

Colouring text

The 5th parameter can be true or false according to whether you want the residue text coloured.  Displaying the residue
boxes and colouring the text will give the impression of no text displayed at all but will slow the display down
considerably.
 

Defining sequences to be included in the consensus

Defining the sequences for the consensus calculation is done in much the same way as for the colour grouping.
 

<PARAM name="Consensus" value="1,4,6-10">

You can also select all the sequences by

<PARAM name="Consensus" value = "*">

 
Defining the SRS server

This is done using the srsServer parameter as follows

<PARAM name="srsServer" value="www.sanger.ac.uk/srs5bin/cgi-bin/">

n.b. the value does NOT have http:// at the front and is otherwise the URL of the SRS program wgetz

(If you are using Netscape you MUST have this server the same as both your mail server and the applet server)
 

Defining the SRS database

Use the database parameter to change your srs database (default is swall).

<PARAM name="database" value="pfamseq">

Defining sequences in html

If you want to you can submit an alignment in the html as follows
 

<PARAM name="numseqs" value=3>

<PARAM name="seq1" value="ALSDKFHLKRHELKHASLDKHLKHR">

<PARAM name="seq2" value="ALSDKFHLKRHE-----LDKHLKHR">

<PARAM name="seq3" value="ALSDKFHLKRHELKHALKFL-HLKHR">

Optionally you can supply parameters that define the ids for each of the sequences as follows:

<PARAM name="id1" value="LCAT_HUMAN">

<PARAM name="id2" value="LCAT_YEAST">

<PARAM name="id3" value="LCAT_MOUSE">

If you don't supply ids then the defaults will be Seq_1 Seq_2 etc.

The Edit menu

• Selecting sequences
• Selecting residues
• Inserting/deleting gaps
• Copying/moving  sequences
• Deleting sequences
• Selecting columns
• Trimming the alignment (Removing sequence -> and <- of selected columns)
• Changing the gap insert character
• Removing gapped columns
• Editing sequence groups (Groups...)
• Editing the alignment by group (Group editing mode)

   Selecting sequences

  Select a sequence with the mouse by  clicking on its name in the left hand panel.  If a sequence is selected its ID background will be grey.  Clicking again will deselect the sequence.  You can select/deselect more than one sequence by dragging the mouse over more than one ID.

  Selected sequences are used in the Colour and Calculate menus.  The Consensus option in the Calculate menu only uses the selected sequences in its calculation and will display an error in its status bar if none are selected.  Selecting a colour scheme in the Colour menu will only apply that scheme if any sequences are selected.  If none are selected that colour scheme is applied to all sequences.

  Sequences can also be selected in other displays such as the tree display window and the PCA results window.  If a sequence is selected in one window it will also be selected in all the other windows.
  

   

  Selecting residues

  Select a residue with the mouse and the status bar will change and display the sequence id and number and the residue's position in the sequence not including gaps.  For instance if we have a sequence in the alignment that is composed of residues 58 to 126 of a swissprot entry selecting a residue in that sequence will report back a number from 58 to 126 and not the position in the alignment.
  
   
   
   

  Inserting/deleting gaps
   

  The alignment may have gaps inserted or deleted by selecting a residue with the mouse and dragging that residue left or right.  Gaps are automatically inserted/deleted but non-gap residues can not be overwritten.
  

   

  Copying/Moving sequences

  After selecting some sequences in the ID panel (or another window)  the selected sequences can be used to make a new alignment window.  Choosing the 'Move selected sequences to new alignment'  will create a new jalview window containing those selected sequences and remove them from the current window.  Choosing the 'Copy selected sequences to new alignment'  will create a new jalview window but leave the sequences in the original window.  Be careful when using the move option as there is no undo function yet.
  

   

  Deleting sequences

  Selected sequences can be deleted by choosing the 'Delete selected sequences' option from the Edit menu.  Be careful with this option as there is no undo function and no going back.
   
   

  Selecting columns
  

  Selecting a position in the Id panel using the mouse will select that column in the alignment and all selected columns are marked with a red box.   This is useful for either remembering a position to come back to or for marking a column for any alignment trimming operations you wish to do.  Choosing the 'Deselect all columns' option in the Edit menu clears any previous column selections.
  
   
   

  Trimming the Alignment
  

  It quite often happens that an alignment you have generated from, say, a blast search contains sequences of varying lengths and has quite ragged edges.  This can bias subsequent calculations such as the PCA analysis or any trees made (phylogenetic or otherwise).  Trimming the alignment left and right to even up the edges can be done by first selecting a column where you wish to trim.  Choosing 'Remove sequence <- left of selected columns' will delete all parts of all sequences to the left of that column and renumber the alignment from that column.  Similarly choosing 'Remove sequence -> right of selected columns'  deletes all parts of the alignment to the right of that column.  If there are more than one column selected then the alignment is trimmed to the leftmost or rightmost column selected.

  All columns may be deselected by choosing the 'Deselect all columns' option from the Edit menu.
  

   

  Setting the gap character

  The gap insert character can be changed from - to . and back again by selecting the 'Set gap character to ...' option in the Edit menu.
  

   

  Removing gapped columns

  It can often happen that you need to removed whole columns consisting of gaps to tidy up your alignment.  This can arise after deleting sequences from an alignment that had large insertions.  Selecting this option from the Edit menu will remove all columns in your alignment that consist entirely of '-', ''.' or ' '.
  

   

  Editing sequence groups (Groups...)

  Sequences can be put into groups which have different colouring schemes applied to them.  This is mostly done automatically now via a combination of  selecting sequences in the Id panel and the Colour menu.   Selecting Edit->Groups will bring up a new window with a list of the existing groups in the top left hand pane.  Selecting one of these groups will cause the sequences contained in that group to be displayed in the next pane.  To add or remove sequences from this group select one or more individual sequences and select a button in the top panel.  Any sequences that aren't in any group will be displayed in the top right most panel.  Display properties for the selected group can be changed using the switches in the botton right hand panel.

  To generate a new group or delete a group the buttons in the bottom right hand of the window can be used.

  Tip:  To generate a new group quickly :
   

  • select your sequences in the ID panel of the main alignment window using the mouse.
  • Bring up the Edit->Groups window.
  • Generate a new group by selecting 'Add new group'
  • Add the selected ids to the new group by pressing 'Add selected IDs'
  This can be useful for generating a group for group editing or putting outliers together in one group prior to conservation analysis.
  

   

  Group editing (Edit->Group edit mode)

  When tweaking an alignment by hand it is useful to be able to move a group of sequences together so the relationship between them is preserved.  Setting this mode on and selecting a residue in the sequence and dragging to insert/delete a gap will cause the same insertions/deletions to occur in all sequences in the same group.

  To create a group from selected sequences or to see the available sequence groups refere to the previous entry.   By default when jalview is first started all sequences are in the same group.

Font menu

• Fastdraw mode
• Different font styles
 

Fastdraw mode

The default setting for jalview is for it to be in Fastdraw mode.  There is only one font available (Courier) and the residues appear more closely packed than for non-fastdraw mode.  If you have a slow machine or a very big alignment then this mode will be the fastest.

If Fastdraw is switched off other proportional fonts can be used (Helvetica and Times) and the residues appear more spaced out on the screen.   The screen update time will also be slower (typically by a factor of 3).


 

Different font styles

By selecting different options from the Font menu the font size and style (plain or bold)  can be changed. These font changes apply to all sequences regardless of whether any sequences are selected or not.

 Smaller font sizes (probably < 6) are of most use if the text is switched off  and the coloured residue boxes only are displayed (see view menu).  

View menu

The options in here refer to how each of the residues in the alignment appears.   Selecting any of these options from this menu will affect every sequence in the alignment regardless of whether any sequences are selected.  If you want to apply these  options to only a subset of sequences this is only possible through the Edit->Groups... menu option.

If the redraw speed is too slow for you on your system then turning off the boxes option  and colouring the text black will speed it up considerably.

 

View boxes
If the Boxes option is ticked then the residue will appear with a background colour.   (The exact colour depends on which colour option is selected in the Colour menu.)

 
 

View text

If the Text option is ticked then the text is also drawn.

 
 

Colour text

If the 'Colour text' option is selected in the Colour menu then the text colour is set to the colour scheme set in the colour menu otherwise it is coloured black.  Obviously if you set both the Boxes option and the colour text option you won't see the residue letters clearly.

 
 

Black text

This option draws all the text in black.

 

Colour menu

• Zappo colours
• Taylor colours
• Hydrophobicity colours
• Colouring residues above a percentage identity threshold
• User defined colour schemes
• PID (Percentage Identity) Colours
• BLOSUM62 colours
• View sequence features
 
This sets the colour scheme for the residue boxes and the residue text.  If a colour scheme is selected from this menu it will affect the selected sequences only.   If no sequences are selected then the colour scheme applies to the whole alignment.
 

Zappo Colours

The residues  are coloured according to their physico-chemical properties as follows:

 

 

Taylor Colours

These colours were invented by Willie Taylor and an entertaining description of their birth can be found in Protein Engineering, Vol 10 , 743-746 (1997)
 

Residues	Full name	Colour
V	Valine
I	Isoleucine
L	Leucine
F	Phenylalanine
Y	Tyrosine
W	Tryptophan
H	Histidine
R	Arginine
K	Lysine
N	Asparagine
Q	Glutamine
E	Glutamate
D	Aspartate
S	Serine
T	Threonine
G	Glycine
P	Proline
C	Cysteine

 
 

Hydrophobicity colours

This option colours the residues in the alignment according to the hydrophobicity table of  Kyte, J., and Doolittle, R.F., J. Mol. Biol. 1157, 105-132, 1982.  The most hydrophobic residues according to this table are coloured red and the most hydrophilic ones are coloured blue.  The colours of the in between residues are varying shades of purple according to whereabouts they are on the scale.


 

User Colours

Once a residue colouring scheme has been selected (Zappo, Taylor or hydrophobicity)  the user can modify these colours to their own specification by selecting the Colour->User defined colours option.  Awindow will appear showing the different colours used for that scheme and which residues are assigned to that colour.  The user can now change the colours used and also which residues are assigned to which colour.

 

Choosing a colour (the colour selector)

Underneath the list of colours and residues is a panel where you can select the rgb values of any colour you wish to use.  The user can either move the scrollbars to change the rgb values or type in the values (0-255) in the text boxes.  The new colour will be displayed in the panel to the right of the scrollbars.


 

Changing colours  (the residue panels)

Clicking on the colours assigned to different residues with the left mouse button will cause whichever colour is displayed in the colour selector to appear in that residue panel.  If you wish to modify an existing residue panel colour right clicking that colour will change the the colour selector's colour to the residue panel colour.  The colour selector colour can then be modified.


 

Changing residues

For each colour present in that scheme a list of residues it is applied to appears to its right.  These residues can be moved or deleted or added to.to group the residues in a different way.  For instance you may just want to display the charged residues in one colour and the rest in another to highlight the charged ones or you may want to only colour the cysteines differently from the others.

If you wish to change the residues associated with a colour edit the residue string in the text field and press the 'Apply' button to its right.  If any residues have been deleted from the text field they will be assigned a white colour and appear in the bottom residue panel.  If any residues have been transferred from another colour panel they will be deleted from the old one.  The main jalview alignment window will be automatically updated.

Any modifications of the colour scheme will only apply to sequences that are selected in the main alignment window.  This allows the user to have multiple colour schemes in one alignment.  If no sequences are selected then the colour scheme applies to all sequences.  BEWARE:: there is NO UNDO function.

 

Colouring above a percentage identity threshold

Selecting this option causes the colour scheme to be applied to only those residues that occur in that column more than a certain percentage of the time.  This is also only applied to selected sequences.  For instance selecting the threshold to be 100 will only colour those columns with 100 % identity.  This threshold option can be applied to the Zappo, Taylor, Hydrophobicity and User colour schemes.

This option depends on a consensus calculation having been performed.  If no consensus exists (e.g. after a copy or a clustalw alignment) then no residues are coloured.


 

PID Colours

This depends on the applet having performed a consensus calculation on the alignment.  This is done automatically when the applet first starts up but is not done for subsequent alignment windows.  If the consensus seuqence is calculated again by selecting 'Consensus' from the Calculate menu then the consensus is calculated for the selected sequences only  and all the alignment sequences are coloured according to that consensus.  This can be useful to display how similar all the other sequences in the alignment are compared to one (or a few) selected sequences.

The PID option colours the residues (boxes and/or text) according to the percentage of the residues in each column that agree with the consensus sequence.  Only the residues that agree with the consensus residue for each column are coloured.

 

Percentage agreement	Colour
> 80 %	Mid blue
> 60 %	Light blue
> 40 %	Light grey
<= 40%	White

 

View sequence features

Selecting this option from the Colour menu will bring up the sequence feature console.  If  your srs server is specified correctly (either on the command line or as an applet parameter)  the database entry for each sequence will be read.  For large sequences this could take quite a time.

When the features have finished transferring the features will be displayed on the alignment with different colours for different features.  The colours are as follows
 
 

Sequence feature	Colour
CHAIN	White
DOMAIN	White
TRANSMEM	Dark red
SIGNAL	Cyan
HELIX	Magenta
TURN	Green
SHEET	Yellow
STRAND	Yellow
CARBOHYD	Pink
ACT_SITE	Red
TRANSIT	Orange
VARIANT	Dark orange
BINDING	Blue
DISULFID	Dark yellow
anything else	Light gray

 

When the features have been displayed on the alignment selecting a residue will change the display in the sequence feature console.  The console will display details of any feature that has been selected and underneath a list of all features listed for that sequence.

There are at the moment a few limitations on the sequence feature display:
 

• Jalview is heavily protein biased and can only parse SWISSPROT features.
 
• Your alignment MUST have the correct start end positions in the ids
 
For example:

>HBA_HUMAN/3-45

means the swissprot ID HBA_HUMAN starting at position 3 and ending at position 45.  If your alignment doesn't have the correct start end positions the sequence feature overlay is at best
meaningless.

A good example of the usage of the start-end positions is the Pfam database of protein alignments.
 

If everything is configured correctly (srs server, database and alignment ids) then you should get output like
the following :

SEE IMAGE

The main window  is coloured using all the features in the Pfam pancreatic trypsin inhibitor alignment and the sequence feature console shows details of all features at the selected residue (which is in between 2 disulphide bonds and at the active site in this case).  In the background can be seen the mini web browser showing the contents of a Swissprot entry.

 

Calculate menu

• Consensus sequence
• Sorting sequence by pairwise identity to consensus
• Sorting by tree order
• Sorting by group order
• Removing redundancy of sequences
• Smith-Waterman pairwise alignment
• Principal component analysis
• UPGMA tree using percentage identity distances
• Neighbour joining tree using percentage identity distances
• Conservation
   

  Consensus sequence

  Each residue in the consensus sequence is the most frequent residue in each column of the alignment excluding gap residues ' ','-' and '.' .  You can't access the consensus sequence directly but it is used  in the PID colour scheme.

  When the editor first starts up the consensus sequence is automatically calculated using all the sequences in the alignment and the PID colour scheme is used as default.  If the consensus option is selected again only the currently selected sequences are used to calculate it and all sequences in the alignment are coloured according to that consensus.
  

   

  Sorting sequences

  Once a consensus calculation has been done selecting this option will sort the selected  sequences by their percentage identity to the consensus sequence.  The most similar sequence is put at the top.  If no sequences are selected then redundancy is removed from the whole alignment.
  

   

  Sorting by tree order

  If a UPGMA tree or a neighbour joining tree has been displayed then the main alignment window displays the sequences in the same order as they appear in the tree.  This makes for easier comparison of the tree and the alignment.
  

   

  Sorting by group order

  If the sequences have been grouped either by hand or by selecting a point on the tree then this option will reorder the alignment so all sequences in the same group are together.  The largest group is shown at the top of the alignment and the smallest at the bottom.
  

   

  Removing redundancy

  Selecting this option brings up a window asking you to select a threshold.   If the percentage identity between two sequences exceeds this value one of the sequences (the shorter) is discarded.    The redundancy calculation is done when the Apply button is pressed.  For large numbers of sequences this can take a long time as all pairs have to be compared.
  

   

  Pairwise alignment (Proteins only)

  This calculation is performed on the selected sequences only.  Java is not the fastest language in the world and aligning more than a handful of sequences will take a fair amount of time.

  For each pair of sequences the best global alignment is found using BLOSUM62 as the scoring matrix.  The scores reported are the raw scores.   The sequences are aligned using a dynamic programming technique and using the following gap penalties :

  Gap open        :   12
  Gap extend    :     2

  When you select the pairwise alignment option a new window will come up which will display the alignments in a text format as they are calculated.  Also displayed is information about the alignment such as alignment score, length and percentage identity between the sequences.

  If you want to save that pairwise alignment (it's not in any known format I'm afraid) you can cut and paste it from the text window with the mouse.  You can also press the 'View in alignment editor'  button to bring up another editor window.
  

   

  Principal Component Analysis

  This is a method of clustering sequences based on the method developed by G. Casari, C. Sander and A. Valencia.   Structural Biology volume 2, no. 2, February 1995 .   Extra information can also be found at the SeqSpace server at the EBI.

  The version implemented here only looks at the clustering of whole sequences and not individual positions in the alignment to help identify functional residues.  This is due to memory and time limitations associated with java (you have to diagonalize a much bigger matrix basically).  Plans are afoot to use the CORBA server written by Chris Dodge to do this 'residue space' PCA remotely.

  When the Calculate->Principal component analysis option is selected all the sequences (not just the selected ones) are used in the calculation and for large numbers of sequences this could take quite a time.  When the calculation is finished a new window is displayed showing the projections of the sequences along the 2nd, 3rd and 4th vectors giving a 3dimensional view of how the sequences cluster.

  This 3d view can be rotated by holding the left mouse button down in the PCA window and moving it.    The user can also zoom in and out by using the up and down arrow keys.

  Individual points can be selected using the mouse and selected sequences show up green in the PCA window and the usual grey background/white text in the alignment and tree windows.

  Different eigenvectors can be used to do the projection by changing the selected dimensions in the 3 menus underneath the 3d window.
  

   

  UPGMA tree

  If this option is selected from the Calculate menu then all sequences are used to generate a UPGMA tree.  The pairwise distances used to cluster the sequences are the percentage  mismatch between two sequences.   For a reliable phylogenetic tree I recommend other programs (phylowin, phylip) should be used as they have the speed to use better distance methods and bootstrapping.  Again, plans are afoot for a server to do this and to be able to read in tree files generated by other programs.

  When the tree has been calculated a new window is displayed showing the tree with labels on the leaves showing the sequence ids.   The user can select the ids with the mouse and the selected sequences will also be selected in the alignment window and the PCA window if  that analysis has been calculated.

  Selecting the 'show distances' checkbox will put branch lengths on the branches.  These branch lengths are the percentage mismatch between two nodes.

  Postscript output can be generated for this tree and mailed to you by clicking the Output button.  This will bring up a window asking  you for your email address  and you can set font options and the page orientation.  Clicking the Apply button will generate the postscript and send the email.
  

   

  Neighbour Joining tree

  The distances between sequences for this tree are generated in the same way as for the UPGMA tree.  The method of clustering is the neighbour joining method which doesn't just pick the two closest leaves to cluster together but compensates for long edges by subtracting from the distances the average distance from each leaf to all the others.

  Selection and output options are the same as for the UPGMA tree.
  

   

  Conservation

  This option is based on the AMAS method of multiple sequence alignment analysis (Livingstone C.D. and Barton G.J. (1993), Protein Sequence Alignments: A Strategy for the Hierarchical Analysis of Residue Conservation.CABIOS Vol. 9 No. 6 (745-756)).

  Hierarchical analysis is based on each residue having certain physico-chemical properties listed as follows:

  In brief  go about it like this :
   

  The alignment can first be divided into groups.  This is best done by first creating an average distance tree (Calculate->Average distance tree).  Selecting a position on the tree will cluster the sequences into groups depending on the position selected.  Each group is coloured a different colour which is used for both the ids in the tree and alignment windows and the sequences themselves.  If a PCA window is visible a visual comparison can be made between the clustering based on the tree and the PCA.

  SEE IMAGE  This link provides an example of the output after grouping for Pfam family rnaseH:

  The grouping by tree may not be satisfactory and the user may want to edit the groups (Edit->Groups...) to put any outliers together.

  Before selecting the conservation option change the colour scheme to something sensible (Taylor or hydrophobicity for example).  When the conservation is done the existing colour scheme is modified so that the most conserved columns in each group have the most intense colours and the least conserved are the palest.

  SEE IMAGE

  This link shows the results of first colouring the alignment by hydrophobicity (Colour->by hydrophobicity) then performing conservation analysis (Calculate->Conservation).  Conserved hydrophobic columns are shown with predominately red residues and conserved hydrophilic columns with blue.  The most conserved regions have the brightest colours.

  SEE IMAGE
   
  Here is shown the same conservation but with Taylor colours instead of hydrophobicity (Colour->Taylor).
   

  The conservation analysis is done on each sequence group.  This highlights differences and similarities  in conserved residue properties between groups.

Multiple alignment

• Local alignment using clustalw
• Remote alignment using clustalw at the EBI
 
 

Local Alignment (Unix applications only)

If you have clustalw installed locally this option will align your sequences on your local machine.    For this option to work you have to have clustalw in your path,  called clustalw and write access to the current directory.

When this option is selected a window will appear giving you a message about whether your process is running and the time elapsed since the job was started.    The cancel button will  kill  your process at any time.

The text box below  should show the progress of your job but at the moment doesn't.  I haven't been able to devise a simple way of  displaying stdout as the alignment is progressing but I'm working on this.  The stdout will appear in the xterm you started

When the alignment is finished a new alignment window is created with the aligned sequences in.  No consensus calculation is done on these sequences by default so to see the similarity select Calculate->Consensus.

Due to applet security restrictions this option can only be used from an application.

 

Remote Alignment

This option sends all your sequences to the EBI and aligns them there.  When the completed alignment is sent back to you a new alignment window is created.

The Cancel button will cancel your job and the output is sent back to the text box below as the alignment progresses.  As this application is written in 1.0 java (pretty much) to enable it to be used in older versions of netscape this display is somewhat flickery.  Using the java1.1 textbox things are much improved  but this won't be available until the majority of browsers support full 1.1.

SRS Access

Jalview provides links to SRS for both the display of sequence features   and for viewing sequence entry annotation in a separate window.

SRS server  and database

The default SRS server  and database are srs.ebi.ac.uk  and swall at the EBI.   To change to your own SRS server either use the -srsserver and -database options on the command line (see command line parameters) or  use the

<param name="srsServer"  value="srs.ebi.ac.uk:5000/srs5bin/cgi-bin/">

and

<param name="database" value="swall">

options in the applet version.

Also for entries to be fetched correctly the sequence IDs in your alignment file must be of the right form.

The IDs must be :

HBA_HUMAN/6-20

or just

HBA_HUMAN

where HBA_HUMAN is the sequence ID (not the accession) and the 6 and the 20 refer to the start and end residues of that sequence entry in the alignment.  The start and end positions do not include gaps and are only essential if you wish to display the sequence features in the alignment window.  I urge everyone to include these numbers as it stops embarrassing mistakes when inferring function from annotation of a multi-domain protein.

The application version now allows access to SRS through it's own mini web browser but at present none of the SRS links work (I'm trying to resist rewriting netscape :)

The alignment window

When the alignment window first appears  you will see a central panel with the sequences in, an ID panel on the left with the sequence identifiers in, a menu bar across the top and a status bar along the bottom.  When an alignment is first loaded a consensus calculation is done.  When this is finished the alignment is coloured according to each residue's percentage identity (PID) agreement with the consensus.

The Status Bar

On the left hand side of the status bar are displayed various messages from the program.  Selecting a residue in the alignment will display the sequence ID of the sequence it is from, its position in the alignment and the residue letter you have selected.  Select an ID in the ID panel and the status bar will display the sequence ID name and its position in the alignment. (n.b. residues and sequences are numbered from 0)  Other messages that appear in the left hand part of the status bar are success/failure of alignment loading, consensus calculation etc.

The right hand side of the status bar is mostly for development purposes but displays in milliseconds the time taken for the last redraw of the central sequence panel.

The ID panel

The identifiers of the sequences are shown on the left hand side of the window.
 

Selecting Sequence IDs

When an alignment is loaded none of the sequences are selected and so menu choices like Calculate Consensus or Pairwise alignments won't work.  To select sequences click with the left mouse button on an ID and the backgound will turn grey to show it's selected.  To unselect it click with the mouse button again.   Dragging the mouse over a number of IDs makes it easier to select/deselect a range of IDs.
 

Showing sequence database entries

If you are running jalview as an applet and your sequence IDs are from swissprot then clicking with the right mouse button on an ID will bring up a new browser window with the swissprot entry in.

* From Michele Clamp (michele@ebi.ac.uk), in http://www.es.embnet.org/Doc/jalview/contents.html.