
   BioPerl DocBook ([1]BioPerl)

Bio::Graphics HOWTO

Lincoln Stein

   [2]Cold Spring Harbor Laboratory

     <[3]lstein@cshl.org>

   This document is copyright Lincoln Stein, 2002. It can be copied and
   distributed under the terms of the Perl Artistic License.

   2002-09-01
   Revision History
   Revision 0.2 2003-05-15 lds
   Current as of BioPerl 1.2.2

   Abstract

   This HOWTO describes how to render sequence data graphically in a
   horizontal map. It applies to a variety of situations ranging from
   rendering the feature table of a GenBank entry, to graphing the
   positions and scores of a BLAST search, to rendering a clone map. It
   describes the programmatic interface to the Bio::Graphics module, and
   discusses how to create dynamic web pages using Bio::DB::GFF and the
   gbrowse package.
     _________________________________________________________________

   Table of Contents

   [4]1. Introduction
   [5]2. Preliminaries
   [6]3. Getting Started
   [7]4. Adding a Scale to the Image
   [8]5. Improving the Image
   [9]6. Parsing Real BLAST Output
   [10]7. Rendering Features from a GenBank or EMBL File
   [11]8. A Better Version of the Feature Renderer
   [12]9. Summary

1. Introduction

   This HOWTO describes the Bio::Graphics module, and some of the
   applications that were built on top of it. Bio::Graphics was designed
   to solve the following common problems:
     * You have a list of BLAST hits on a sequence and you want to
       generate a picture that shows where the hits go and what their
       score is.
     * You have a big GenBank file with a complex feature table, and you
       want to render the positions of the genes, repeats, promoters and
       other features.
     * You have a list of ESTs that you've mapped to a genome, and you
       want to show how they align.
     * You have created a clone fingerprint map, and you want to display
       it.

   The Bio::Graphics module was designed to solve these problems. In
   addition, using the Bio::DB::GFF module (part of BioPerl) and the
   gbrowse program (available from http://www.gmod.org) you can create
   interactive web pages to explore your data.

   This document takes you through a few common applications of
   Bio::Graphics in a cookbook fashion.

2. Preliminaries

   Bio::Graphics is dependent on GD, a Perl module for generating
   bitmapped graphics written by the author. GD in turn is dependent on
   libgd, a C library written by Thomas Boutell, formerly also of Cold
   Spring Harbor Laboratory (www.boutell.com/gd). To use Bio::Graphics,
   you must have both these software libraries installed.

   If you are on a Linux system, you might already have GD installed. To
   verify, run the following command:

 % perl -MGD -e 'print $GD::VERSION';


   If the program prints out a version number, you are in luck.
   Otherwise, if you get a "Can't locate GD.pm" error, you'll have to
   install the module. For users of ActiveState Perl this is very easy.
   Just start up the PPM program and issue the command "install GD". For
   users of other versions of Perl, you should go to www.cpan.org,
   download a recent version of the GD module, unpack it, and follow the
   installation directions. You may also need to upgrade to a recent
   version of the libgd C library.

   You may need to upgrade to a recent version of the libgd C library. At
   the time this was written, there were two versions of libgd. libgd
   version 1.8.4 is the stable version, and corresponds to GD version
   1.43. libgd version 2.0.1 is the beta version; although it has many
   cool features, it also has a few known bugs (which Bio::Graphics works
   around). If you use libgd 2.0.1 or higher, be sure it matches GD
   version 2.0.1 or higher.

   You will also need to install the Text::Shellwords module, which is
   available from CPAN.

3. Getting Started

   All the code examples and BLAST input files we'll use are available in
   the doc/howto/examples/graphics directory in the BioPerl package.

   Our first example will be rendering a table of BLAST hits on a
   sequence that is exactly 1000 residues long. For now, we're ignoring
   finicky little details like HSPs, and assume that each hit is a single
   span from start to end. Also, we'll be using the BLAST score rather
   than P or E value. Later on, we'll switch to using real BLAST output
   parsed by the Bio::SearchIO module, but for now, our table looks like
   this:

# hit           score   start   end
hsHOX3          381     2       200
scHOX3          210     2       210
xlHOX3          800     2       200
hsHOX2          1000    380     921
scHOX2          812     402     972
xlHOX2          1200    400     970
BUM             400     300     620
PRES1           127     310     700

   Figure 1. Simple blast hit file (data1.txt)

   Our first attempt to parse and render this file looks like this:

   Example 1. Rendering the simple blast hit file (render_blast1.pl)
  0  #!/usr/bin/perl

  1  # This is code example 1 in the Graphics-HOWTO
  2  use strict;
  3  use Bio::Graphics;
  4  use Bio::SeqFeature::Generic;

  5  my $panel = Bio::Graphics::Panel->new(-length => 1000,-width  => 800);
  6  my $track = $panel->add_track(-glyph => 'generic',-label  => 1);

  7  while (<>) { # read blast file
  8    chomp;
  9    next if /^\#/;  # ignore comments
 10    my($name,$score,$start,$end) = split /\t+/;
 11    my $feature = Bio::SeqFeature::Generic->new(-display_name=>$name,-score=
>$score,
 12                                                -start=>$start,-end=>$end);
 13    $track->add_feature($feature);
 14  }

 15  print $panel->png;

   The script begins by loading the Bio::Graphics module (line 3), which
   in turn brings in a number of other modules that we'll use later. We
   also load Bio::SeqFeature::Generic in order to create a series of
   Bio::SeqFeatureI objects for rendering. We then create a
   Bio::Graphics::Panel object by calling its new() method, specifying
   that the panel is to correspond to a sequence that is 1000 nucleotides
   long, and has a physical width of 800 pixels (line 5). The Panel can
   contain multiple horizontal tracks, each of which has its own way of
   rendering features (called a "glyph"), color, labeling convention, and
   so forth. In this simple example, we create a single track by calling
   the panel object's add_track() method (line 6), specify a glyph type
   of "generic", and ask that the objects in the track be labeled by
   providing a true value to the -label argument. This gives us a track
   object that we can add our hits to.

   We're now ready to render the blast hit file. We loop through it (line
   7-14), stripping off the comments, and parsing out the name, score and
   range (line 10). We now need a Bio::SeqFeatureI object to place in the
   track. The easiest way to do this is to create a
   Bio::SeqFeature::Generic object, which is similar to Bio::PrimarySeq,
   except that it provides a way of attaching start and end positions to
   the sequence, as well as such nebulous but useful attributes as the
   "score" and "source". The Bio::SeqFeature::Generic->new() method,
   invoked in line 11, takes arguments corresponding to the name of each
   hit, its start and end coordinates, and its score.

   After creating the feature object, we add it to the track by calling
   the track's add_feature() method (line 13).

   After processing all the hits, we call the panel's png() method to
   render them and convert it into a Portable Network Graphics file, the
   contents of which are printed to standard output. We can now view the
   result by piping it to our favorite image display program.
   [Important]
   Important

   If you are on a Windows platform, you need to put STDOUT into binary
   mode so that the PNG file does not go through Window's carriage
   return/linefeed transformations. Before the final print statement, put
   the statement "binmode(STDOUT)".

   This advice also applies to certain versions of RedHat, which ship
   with a patched (and possibly broken) version of Perl.

% render_blast1.pl data1.txt | display -

   Rendering BLAST hits

   Figure 2. Rendering BLAST hits

   Users of operating systems that don't support pipes can simply
   redirect the output to a file and view it in their favorite image
   program.

4. Adding a Scale to the Image

   This is all very nice, but it's missing two essential components:
     * It doesn't have a scale.
     * It doesn't distinguish between hits with different scores.

   Example 2 fixes these problems

   Example 2. Rendering the blast hit file with scores and scale
  0  #!/usr/bin/perl

  1  # This is code example 2 in the Graphics-HOWTO
  2  use strict;
  3  use lib '/home/lstein/projects/bioperl-live';
  4  use Bio::Graphics;
  5  use Bio::SeqFeature::Generic;

  6  my $panel = Bio::Graphics::Panel->new(-length => 1000,
  7                                        -width  => 800,
  8                                        -pad_left => 10,
  9                                        -pad_right => 10,
 10                                       );
 11  my $full_length = Bio::SeqFeature::Generic->new(-start=>1,-end=>1000);
 12  $panel->add_track($full_length,
 13                    -glyph   => 'arrow',
 14                    -tick    => 2,
 15                    -fgcolor => 'black',
 16                    -double  => 1,
 17                   );

 18  my $track = $panel->add_track(-glyph => 'graded_segments',
 19                                -label  => 1,
 20                                -bgcolor => 'blue',
 21                                -min_score => 0,
 22                                -max_score => 1000);

 23  while (<>) { # read blast file
 24    chomp;
 25    next if /^\#/;  # ignore comments
 26    my($name,$score,$start,$end) = split /\t+/;
 27    my $feature = Bio::SeqFeature::Generic->new(-display_name=>$name,-score=
>$score,
 28                                                -start=>$start,-end=>$end);
 29    $track->add_feature($feature);
 30  }

 31  print $panel->png;

   There are several changes to look at. The first is minor. We'd like to
   put a boundary around the left and right edges of the image so that
   the features don't bump up against the margin, so we specify a 10
   pixel leeway with the -pad_left and -pad_right arguments in lines 8
   and 9.

   The next change is more subtle. We want to draw a scale all the way
   across the image. To do this, we create a track to contain the scale,
   and a feature that spans the track from the start to the end. Line 11
   creates the feature, giving its start and end coordinates. Lines 12-17
   create a new track containing this feature. Unlike the previous
   example, in which we created the track first and then added features
   one at a time with add_feature(), we show here how to add feature(s)
   directly in the call to add_track(). If the first argument to
   add_track is either a single feature or a feature array ref, then
   add_track() will automatically incorporate the feature(s) into the
   track in a single efficient step. The remainder of the arguments
   configure the track as before. The -glyph argument says to use the
   "arrow" glyph. The -tick argument indicates that the arrow should
   contain tick marks, and that both major and minor ticks should be
   shown (tick type of "2"). We set the foreground color to black, and
   request that arrows should be placed at both ends (-double =>1).

   [[13]1]

   In lines 18-22, we get a bit fancier with the blast hit track. Now,
   instead of creating a generic glyph, we use the "graded_segments"
   glyph. This glyph takes the specified background color for the
   feature, and either darkens or lightens it according to its score. We
   specify the base background color (-bgcolor => 'blue'), and the
   minimum and maximum scores to scale to (-min_score and -max_score).
   (You may need to experiment with the min and max scores in order to
   get the glyph to scale the colors the way you want.) The remainder of
   the program is the same.

   When we run the modified script, we get this image.

   The improved image

   Figure 3. The improved image
   [Important]
   Important

   Remember that if you are on a Windows platform, you need to put STDOUT
   into binary mode so that the PNG file does not go through Window's
   carriage return/linefeed transformations. Before the final print
   statement, write binmode(STDOUT).

5. Improving the Image

   Before we move into displaying gapped alignments, let's tweak the
   image slightly so that higher scoring hits appear at the top of the
   image, and the score itself is printed in red underneath each hit. The
   changes are shown in Example 3.

   Example 3. Rendering the blast hit file with scores and scale
  0  #!/usr/bin/perl

  1  # This is code example 3 in the Graphics-HOWTO
  2  use strict;
  3  use lib '/home/lstein/projects/bioperl-live';
  4  use Bio::Graphics;
  5  use Bio::SeqFeature::Generic;

  6  my $panel = Bio::Graphics::Panel->new(-length => 1000,
  7                                        -width  => 800,
  8                                        -pad_left => 10,
  9                                        -pad_right => 10,
 10                                       );
 11  my $full_length = Bio::SeqFeature::Generic->new(-start=>1,-end=>1000);
 12  $panel->add_track($full_length,
 13                    -glyph   => 'arrow',
 14                    -tick    => 2,
 15                    -fgcolor => 'black',
 16                    -double  => 1,
 17                   );

 18  my $track = $panel->add_track(-glyph => 'graded_segments',
 19                                -label  => 1,
 20                                -bgcolor => 'blue',
 21                                -min_score => 0,
 22                                -max_score => 1000,
 23                                -font2color     => 'red',
 24                                -sort_order     => 'high_score',
 25                                -description => sub {
 26                                  my $feature = shift;
 27                                  my $score   = $feature->score;
 28                                  return "score=$score";
 29                                 });

 30  while (<>) { # read blast file
 31    chomp;
 32    next if /^\#/;  # ignore comments
 33    my($name,$score,$start,$end) = split /\t+/;
 34    my $feature = Bio::SeqFeature::Generic->new(-score=>$score,
 35                                                -display_name=>$name,
 36                                                -start=>$start,-end=>$end);
 37    $track->add_feature($feature);
 38  }

 39  print $panel->png;

   There are two changes to look at. The first appears in line 24, where
   we pass the -sort_order option to the call that creates the blast hit
   track. -sort_order changes the way that features sort from top to
   bottom, and will accept a number of prepackaged sort orders or a
   coderef for custom sorting. In this case, we pass a prepackaged sort
   order of high_score, which sorts the hits from top to bottom in
   reverse order of their score.

   The second change is more complicated, and involves the -description
   option that appears in the add_track() call on lines 25-28. The value
   of -description will be printed beneath each feature. We could pass
   -description a constant string, but that would simply print the same
   string under each feature. Instead we pass -description a code
   reference to a subroutine that will be invoked while the picture is
   being rendered. This subroutine will be passed the current feature,
   and must return the string to use as the value of the description. In
   our code, we simply fetch out the BLAST hit's score using its score()
   method, and incorporate that into the description string.
   [Tip]
   Tip

   The ability to use a code reference as a configuration option isn't
   unique to -description. In fact, you can use a code reference for any
   of the options passed to add_track().

   Another minor change is the use of -font2color in line 23. This simply
   sets the color used for the description strings. Figure 3 shows the
   effect of these changes.

   The image with descriptions and sorted hits

   Figure 4. The image with descriptions and sorted hits

6. Parsing Real BLAST Output

   From here it's just a small step to writing a general purpose utility
   that will read a BLAST file, parse its output, and output a picture.
   The key is to use the Bio::SearchIO infrastructure because it produces
   Bio::SeqFeatureI similarity hits that can be rendered directly by
   Bio::Graphics.

   Code example 4 shows the new utility.

   Example 4. Parsing and Rendering a Real BLAST File with Bio::SearchIO
  0  #!/usr/bin/perl

  1  # This is code example 4 in the Graphics-HOWTO
  2  use strict;
  3  use lib "$ENV{HOME}/projects/bioperl-live";
  4  use Bio::Graphics;
  5  use Bio::SearchIO;

  6  my $file = shift or die "Usage: render_blast4.pl <blast file>\n";

  7  my $searchio = Bio::SearchIO->new(-file   => $file,
  8                                       -format => 'blast') or die "parse fai
led";


  9  my $result = $searchio->next_result() or die "no result";

 10  my $panel = Bio::Graphics::Panel->new(-length    => $result->query_length,
 11                                        -width     => 800,
 12                                        -pad_left  => 10,
 13                                        -pad_right => 10,
 14                                       );

 15  my $full_length = Bio::SeqFeature::Generic->new(-start=>1,-end=>$result->q
uery_length,
 16                                                  -display_name=>$result->qu
ery_name);
 17  $panel->add_track($full_length,
 18                    -glyph   => 'arrow',
 19                    -tick    => 2,
 20                    -fgcolor => 'black',
 21                    -double  => 1,
 22                    -label   => 1,
 23                   );

 24  my $track = $panel->add_track(-glyph       => 'graded_segments',
 25                                -label       => 1,
 26                                -connector   => 'dashed',
 27                                -bgcolor     => 'blue',
 28                                -font2color  => 'red',
 29                                -sort_order  => 'high_score',
 30                                -description => sub {
 31                                  my $feature = shift;
 32                                  return unless $feature->has_tag('descripti
on');
 33                                  my ($description) = $feature->each_tag_val
ue('description');
 34                                  my $score = $feature->score;
 35                                  "$description, score=$score";
 36                                 });

 37  while( my $hit = $result->next_hit ) {
 38    next unless $hit->significance < 1E-20;
 39    my $feature = Bio::SeqFeature::Generic->new(-score   => $hit->raw_score,
 40                                                -display_name => $hit->name,
 41                                                -tag     => {
 42                                                             description =>
$hit->description
 43                                                            },
 44                                               );
 45    while( my $hsp = $hit->next_hsp ) {
 46      $feature->add_sub_SeqFeature($hsp,'EXPAND');
 47    }

 48    $track->add_feature($feature);
 49  }

 50  print $panel->png;

   In lines 6-8 we read the name of the file that contains the BLAST
   results from the command line, and pass it to Bio::SearchIO->new(),
   returning a Bio::SearchIO object. We read a single result from the
   searchIO object (line 9). This assumes that the BLAST output file
   contains a single run of BLAST only.

   We then initialize the panel and tracks as before. The only change
   here is in lines 24-36, where we create the track for the BLAST hits.
   The -description option has now been enhanced to create a line of text
   that incorporates the "description" tag from the feature object as
   well as its similarity score. There's also a slight change in line 26,
   where we introduce the -connector option. This allows us to configure
   a line that connects the segments of a discontinuous feature, such as
   the HSPs in a BLAST hit. In this case, we asked the rendering engine
   to produce a dashed connector line.

   The remainder of the script retrieves each of the hits from the BLAST
   file, creates a Feature object representing the hit, and then
   retrieves each HSP and incorporates it into the feature. Line 37
   begins a while() loop that retrieves each of the similarity hits in
   turn. We filter the hit by its significance, throwing out any that
   have an expectation value greater than 1E-20 (you will have to adjust
   this in your own utilities). We then use the information in the hit to
   construct a Bio::SeqFeature::Generic object (lines 39-44). Notice how
   the name of the hit and the score are used to initialize the feature,
   and how the description is turned into a tag named "description."

   The start and end bounds of the hit are determined by the union of its
   HSPs. We loop through each of the hit's HSPs by calling its next_hsp()
   method, and add each HSP to the newly-created hit feature by calling
   the feature's add_sub_SeqFeature() method (line 46). The EXPAND
   parameter instructs the feature to expand its start and end
   coordinates to enclose the added subfeature.

   Once all the HSPs are added to the feature, we insert the feature into
   the track as before using the track's add_feature() function.

   Figure 4 shows the output from a sample BLAST hit file.

   Output from the BLAST parsing and rendering script

   Figure 5. Output from the BLAST parsing and rendering script

   The next section will demonstrate how to parse and display feature
   tables from GenBank and EMBL.
   [Important]
   Important

   Remember that if you are on a Windows platform, you need to put STDOUT
   into binary mode so that the PNG file does not go through Window's
   carriage return/linefeed transformations. Before the final print
   statement, write binmode(STDOUT).

7. Rendering Features from a GenBank or EMBL File

   With Bio::Graphics you can render the feature table of a GenBank or
   EMBL file quite easily. The trick is to use Bio::SeqIO to generate a
   set of Bio::SeqFeatureI objects, and to use those features to populate
   tracks (see the [14]Feature-Annotation HOWTO for more information on
   features). The documentation for each of the individual. For
   simplicity's sake, we will sort each feature by its primary tag (such
   as "exon") and create a new track for each tag type.

   Code example 5 shows the code for rendering an EMBL or GenBank entry.

   Example 5. The embl2picture.pl script turns any EMBL or GenBank entry
   into a graphical rendering
  0  #!/usr/bin/perl

  1  # file: embl2picture.pl
  2  # This is code example 5 in the Graphics-HOWTO
  3  # Author: Lincoln Stein

  4  use strict;
  5  use lib "$ENV{HOME}/projects/bioperl-live";
  6  use Bio::Graphics;
  7  use Bio::SeqIO;
  8  use Bio::SeqFeature::Generic;

  9  my $file = shift                       or die "provide a sequence file as
the argument";
 10  my $io = Bio::SeqIO->new(-file=>$file) or die "couldn't create Bio::SeqIO"
;
 11  my $seq = $io->next_seq                or die "couldn't find a sequence in
 the file";
 12  my $wholeseq = Bio::SeqFeature::Generic->new(-start=>1,-end=>$seq->length,
 13                                               -display_name=>$seq->display_
name);

 14  my @features = $seq->all_SeqFeatures;

 15  # partition features by their primary tags
 16  my %sorted_features;
 17  for my $f (@features) {
 18    my $tag = $f->primary_tag;
 19    push @{$sorted_features{$tag}},$f;
 20  }

 21  my $panel = Bio::Graphics::Panel->new(
 22                                        -length    => $seq->length,
 23                                        -key_style => 'between',
 24                                        -width     => 800,
 25                                        -pad_left  => 10,
 26                                        -pad_right => 10,
 27                                        );
 28  $panel->add_track($wholeseq,
 29                    -glyph => 'arrow',
 30                    -bump => 0,
 31                    -double=>1,
 32                    -tick => 2);

 33  $panel->add_track($wholeseq,
 34                    -glyph  => 'generic',
 35                    -bgcolor => 'blue',
 36                    -label  => 1,
 37                   );

 38  # general case
 39  my @colors = qw(cyan orange blue purple green chartreuse magenta yellow aq
ua);
 40  my $idx    = 0;
 41  for my $tag (sort keys %sorted_features) {
 42    my $features = $sorted_features{$tag};
 43    $panel->add_track($features,
 44                      -glyph    =>  'generic',
 45                      -bgcolor  =>  $colors[$idx++ % @colors],
 46                      -fgcolor  => 'black',
 47                      -font2color => 'red',
 48                      -key      => "${tag}s",
 49                      -bump     => +1,
 50                      -height   => 8,
 51                      -label    => 1,
 52                      -description => 1,
 53                     );
 54  }

 55  print $panel->png;
 56  exit 0;

   The way this script works is simple. After the library load preamble,
   the script reads the name of the GenBank or EMBL file from the command
   line (line 8). It passes the filename to Bio::SeqIO's new() method,
   and reads the first sequence object from it (lines 9-11). If anything
   goes wrong, the script dies with an error message.

   The returned object is a Bio::SeqI object, which has a length but no
   defined start or end coordinates. We would like to create a drawable
   Bio::SeqFeatureI object to use for the scale, so we generate a new
   Bio::SeqFeature::Generic object that goes from a start of 1 to the
   length of the sequence. (lines 12-13).

   The script reads the features from the sequence object by calling
   all_SeqFeatures(), and then sorts each feature by its primary tag into
   a hash of array references named %sorted_features (lines 14-20).

   Next, we create the Bio::Graphics::Panel object (lines 21-27). As in
   previous examples, we specify the width of the image, as well as some
   extra white space to pad out the left and right borders.

   We now add two tracks, one for the scale (lines 28-32) and the other
   for the sequence as a whole (33-37). As in the earlier examples, we
   pass add_track() the sequence object as the first argument before the
   options so that the object is incorporated into the track immediately.

   We are now ready to create a track for each feature type. In order to
   distinguish the tracks by color, we initialize an array of 9 color
   names and simply cycle through them (lines 39-54). For each feature
   tag, we retrieve the corresponding list of features from
   %sorted_features (line 42) and create a track for it using the
   "generic" glyph and the next color in the list (lines 43-53). We set
   the -label and -description options to the value "1". This signals
   Bio::Graphics that it should do the best it can to choose useful label
   and description values on its own.

   After adding all the feature types, we call the panel's png() method
   to generate a graphic file, which we print to STDOUT. If we are on a
   Windows platform, we would have to include binmode(STDOUT) prior to
   this statement in order to avoid Windows textmode carriage
   return/linefeed transformations.

   Figure 5 shows an example of the output of this script.

   The embl2picture.pl script

   Figure 6. The embl2picture.pl script

8. A Better Version of the Feature Renderer

   The previous example's rendering has numerous deficiencies. For one
   thing, there are no lines connecting the various CDS rectangles in the
   CDS track to show how they are organized into a spliced transcript.
   For another, the repetition of the source tag "EMBL/GenBank/SwissProt"
   is not particularly illuminating.

   However, it's quite easy to customize the display, making the script
   into a generally useful utility. The revised code is shown in example
   6.

   Example 6. The embl2picture.pl script turns any EMBL or GenBank entry
   into a graphical rendering
  0  #!/usr/bin/perl

  1  # file: embl2picture.pl
  2  # This is code example 6 in the Graphics-HOWTO
  3  # Author: Lincoln Stein

  4  use strict;
  5  use lib "$ENV{HOME}/projects/bioperl-live";
  6  use Bio::Graphics;
  7  use Bio::SeqIO;

  8  use constant USAGE =><<END;
  9  Usage: $0 <file>
 10     Render a GenBank/EMBL entry into drawable form.
 11     Return as a GIF or PNG image on standard output.

 12     File must be in embl, genbank, or another SeqIO-
 13     recognized format.  Only the first entry will be
 14     rendered.

 15  Example to try:
 16     embl2picture.pl factor7.embl | display -

 17  END

 18  my $file = shift                       or die USAGE;
 19  my $io = Bio::SeqIO->new(-file=>$file) or die USAGE;
 20  my $seq = $io->next_seq                or die USAGE;
 21  my $wholeseq = Bio::SeqFeature::Generic->new(-start=>1,-end=>$seq->length,
 22                                               -display_name=>$seq->display_
name);

 23  my @features = $seq->all_SeqFeatures;

 24  # sort features by their primary tags
 25  my %sorted_features;
 26  for my $f (@features) {
 27    my $tag = $f->primary_tag;
 28    push @{$sorted_features{$tag}},$f;
 29  }

 30  my $panel = Bio::Graphics::Panel->new(
 31                                        -length    => $seq->length,
 32                                        -key_style => 'between',
 33                                        -width     => 800,
 34                                        -pad_left  => 10,
 35                                        -pad_right => 10,
 36                                        );
 37  $panel->add_track($wholeseq,
 38                    -glyph => 'arrow',
 39                    -bump => 0,
 40                    -double=>1,
 41                    -tick => 2);

 42  $panel->add_track($wholeseq,
 43                    -glyph  => 'generic',
 44                    -bgcolor => 'blue',
 45                    -label  => 1,
 46                   );

 47  # special cases
 48  if ($sorted_features{CDS}) {
 49    $panel->add_track($sorted_features{CDS},
 50                      -glyph      => 'transcript2',
 51                      -bgcolor    => 'orange',
 52                      -fgcolor    => 'black',
 53                      -font2color => 'red',
 54                      -key        => 'CDS',
 55                      -bump       =>  +1,
 56                      -height     =>  12,
 57                      -label      => \&gene_label,
 58                      -description=> \&gene_description,
 59                     );
 60    delete $sorted_features{'CDS'};
 61  }

 62  if ($sorted_features{tRNA}) {
 63    $panel->add_track($sorted_features{tRNA},
 64                      -glyph     =>  'transcript2',
 65                      -bgcolor   =>  'red',
 66                      -fgcolor   =>  'black',
 67                      -font2color => 'red',
 68                      -key       => 'tRNAs',
 69                      -bump      =>  +1,
 70                      -height    =>  12,
 71                      -label     => \&gene_label,
 72                     );
 73    delete $sorted_features{tRNA};
 74  }

 75  # general case
 76  my @colors = qw(cyan orange blue purple green chartreuse magenta yellow aq
ua);
 77  my $idx    = 0;
 78  for my $tag (sort keys %sorted_features) {
 79    my $features = $sorted_features{$tag};
 80    $panel->add_track($features,
 81                      -glyph    =>  'generic',
 82                      -bgcolor  =>  $colors[$idx++ % @colors],
 83                      -fgcolor  => 'black',
 84                      -font2color => 'red',
 85                      -key      => "${tag}s",
 86                      -bump     => +1,
 87                      -height   => 8,
 88                      -description => \&generic_description
 89                     );
 90  }

 91  print $panel->png;
 92  exit 0;

 93  sub gene_label {
 94    my $feature = shift;
 95    my @notes;
 96    foreach (qw(product gene)) {
 97      next unless $feature->has_tag($_);
 98      @notes = $feature->each_tag_value($_);
 99      last;
100    }
101    $notes[0];
102  }

103  sub gene_description {
104    my $feature = shift;
105    my @notes;
106    foreach (qw(note)) {
107      next unless $feature->has_tag($_);
108      @notes = $feature->each_tag_value($_);
109      last;
110    }
111    return unless @notes;
112    substr($notes[0],30) = '...' if length $notes[0] > 30;
113    $notes[0];
114  }

115  sub generic_description {
116    my $feature = shift;
117    my $description;
118    foreach ($feature->all_tags) {
119      my @values = $feature->each_tag_value($_);
120      $description .= $_ eq 'note' ? "@values" : "$_=@values; ";
121    }
122    $description =~ s/; $//; # get rid of last
123    $description;
124  }

   At 124 lines, this is the longest example in this HOWTO, but the
   changes are straightforward. The major difference occurs in lines
   47-61 and 62-74, where we handle two special cases: "CDS" records and
   "tRNAs". For these two feature types we would like to draw the
   features like genes using the "transcript2" glyph. This glyph draws
   inverted V's for introns, if there are any, and will turn the last (or
   only) exon into an arrow to indicate the direction of transcription.

   First we look to see whether there are any features with the primary
   tag of "CDS" (lines 47-61). If so, we create a track for them using
   the desired glyph. Line 49 shows how to add several features to a
   track at creation time. If the first argument to add_track() is an
   array reference, all the features contained in the array will be
   incorporated into the track. We provide custom code references for the
   -label and -description options. As we shall see later, the
   subroutines these code references point to are responsible for
   extracting names and descriptions for the coding regions. After we
   handle this special case, we remove the CDS feature type from the
   %sorted_features array.

   We do the same thing for tRNA features, but with a different color
   scheme (lines 62-74).

   Having dealt with the special cases, we render the remaining feature
   types using the same code we used earlier. The only change is that
   instead of allowing Bio::Graphics::Panel to guess at the description
   from the feature's source tag, we use the -description option to point
   to a subroutine that will generate more informative description
   strings.

   The gene_label() (lines 93-102) and gene_description() (lines 103-114)
   subroutines are simple. The first one searches the feature for the
   tags "product" and/or "gene" and uses the first one it finds as the
   label for the feature. The gene_description() subroutine is similar,
   except that it returns the value of the first tag named "note". If the
   description is over 30 characters long, it is truncated.

   The generic_description() (lines 115-124) is invoked to generate
   descriptions of all non-gene features. We simply concatenate together
   the names and values of tags. For example the entry:
   source          1..12850
                   /db_xref="taxon:9606"
                   /organism="Homo sapiens"
                   /map="13q34"


   will be turned into the description string "db_xref=taxon:9606;
   organism=Homo Sapiens; map=13q34".

   After adding all the feature types, we call the panel's png() method
   to generate a graphic file, which we print to STDOUT.

   Figure 6 shows an example of the output of this script.

   The embl2picture.pl script

   Figure 7. The embl2picture.pl script

9. Summary

   In summary, we have seen how to use the Bio::Graphics module to
   generate representations of sequence features as horizontal maps. We
   applied these techniques to two common problems: rendering the output
   of a BLAST run, and rendering the feature table of a GenBank/EMBL
   entry.

   The graphics module is quite flexible. In addition to the options that
   we have seen, there are glyphs for generating point-like features such
   as SNPs, specialized glyphs that draw GC content and open reading
   frames, and glyphs that generate histograms, bar charts and other
   types of graphs. Bio::Graphics has been used to represent physical
   (clone) maps, radiation hybrid maps, EST clusters, cytogenetic maps,
   restriction maps, and much more.

   Although we haven't shown it, Bio::Graphics provides support for
   generating HTML image maps. The [15]Generic Genome Browser uses this
   facility to generate clickable, browsable images of the genome from a
   variety of genome databases.

   Another application you should investigate is the render_sequence.pl
   script. This script uses the BioFetch interface to fetch
   GenBank/EMBL/SwissProt entries dynamically from the web before
   rendering them into PNG images.

   Finally, if you find yourself constantly tweaking the graphic options,
   you might be interested in Bio::Graphics::FeatureFile, a utility
   module for interpreting and rendering a simple tab-delimited format
   for sequence features. feature_draw.PLS is a Perl script built on top
   of this module, which you can find in the scripts/graphics directory
   in the Bioperl distribution.
   _________________________________________________________________

   [[16]1] Obtain the list of glyphs by running perldoc on
   Bio::Graphics::Glyph. Obtain a description of the glyph options by
   running perldoc on individual glyphs, for example "perldoc
   Bio::Graphics::Glyph::arrow."

   BioPerl DocBook ([17]BioPerl)

References

   1. http://bioperl.org/
   2. http://www.cshl.org/
   3. mailto:lstein@cshl.org
   4. file://localhost/Users/bosborne/bioperl-live/doc/howto/xml/Graphics-HOWTO.html#intro
   5. file://localhost/Users/bosborne/bioperl-live/doc/howto/xml/Graphics-HOWTO.html#prelim
   6. file://localhost/Users/bosborne/bioperl-live/doc/howto/xml/Graphics-HOWTO.html#gettingStarted
   7. file://localhost/Users/bosborne/bioperl-live/doc/howto/xml/Graphics-HOWTO.html#addingscale
   8. file://localhost/Users/bosborne/bioperl-live/doc/howto/xml/Graphics-HOWTO.html#improving
   9. file://localhost/Users/bosborne/bioperl-live/doc/howto/xml/Graphics-HOWTO.html#parsing-blast
  10. file://localhost/Users/bosborne/bioperl-live/doc/howto/xml/Graphics-HOWTO.html#rendering-features
  11. file://localhost/Users/bosborne/bioperl-live/doc/howto/xml/Graphics-HOWTO.html#rendering-features-better
  12. file://localhost/Users/bosborne/bioperl-live/doc/howto/xml/Graphics-HOWTO.html#summary
  13. file://localhost/Users/bosborne/bioperl-live/doc/howto/xml/Graphics-HOWTO.html#ftn.d4e94
  14. http://bioperl.org/HOWTOs/html/Feature-Annotation.html
  15. http://www.gmod.org/
  16. file://localhost/Users/bosborne/bioperl-live/doc/howto/xml/Graphics-HOWTO.html#d4e94
  17. http://bioperl.org/
