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CONTENTS

A. CREATING NEW DATABASES FROM SCRATCH
B. ADDING A NEW DATABASE TO THE BROWSER
C. GENERATING HISTOGRAMS
D. INTERNATIONALIZATION
E. AUTHENTICATION AND AUTHORIZATION
F. DISPLAYING GENETIC AND RH MAPS
G. CHANGING THE LOCATION OF THE CONFIGURATION FILES
H. USING DAS (DISTRIBUTED ANNOTATION SYSTEM) DATABASES
I. THE BioMOBY BROWSER
J. BIOMOBY SERVICES
K. FILTERING SEARCH RESULTS
L. INVOKING GBROWSE URLs
M. FURTHER INFORMATION

A. CREATING NEW DATABASES FROM SCRATCH ^

This section describes how to create new annotation databases from scratch.

A1. The GFF file format

GBrowse is based around the GFF file format, which stands for "Gene Finding Format" and was invented at the Sanger Centre. The GFF format is a flat tab-delimited file, each line of which corresponds to an annotation, or feature. Each line has nine columns and looks like this:

 Chr1  curated  CDS 365647  365963  .  +  1  Transcript "R119.7"

The 9 columns are as follows:

1 reference sequence

This is the ID of the sequence that is used to establish the coordinate system of the annotation. In the example above, the reference sequence is "Chr1".

2 source

The source of the annotation. This field describes how the annotation was derived. In the example above, the source is "curated" to indicate that the feature is the result of human curation. The names and versions of software programs are often used for the source field, as in "tRNAScan-SE/1.2".

3 method

The annotation method. This field describes the type of the annotation, such as "CDS". Together the method and source describe the annotation type.

4 start position

The start of the annotation relative to the reference sequence.

5 stop position

The stop of the annotation relative to the reference sequence. Start is always less than or equal to stop.

6 score

For annotations that are associated with a numeric score (for example, a sequence similarity), this field describes the score. The score units are completely unspecified, but for sequence similarities, it is typically percent identity. Annotations that don't have a score can use "."

7 strand

For those annotations which are strand-specific, this field is the strand on which the annotation resides. It is "+" for the forward strand, "-" for the reverse strand, or "." for annotations that are not stranded.

8 phase

For annotations that are linked to proteins, this field describes the phase of the annotation on the codons. It is a number from 0 to 2, or "." for features that have no phase.

9 group

GFF provides a simple way of generating annotation hierarchies ("is composed of" relationships) by providing a group field. The group field contains the class and ID of an annotation which is the logical parent of the current one. In the example given above, the group is the Transcript named "R119.7".

The group field is also used to store information about the target of sequence similarity hits, and miscellaneous notes. See the next section for a description of how to describe similarity targets.

The sequences used to establish the coordinate system for annotations can correspond to sequenced clones, clone fragments, contigs or super-contigs.

In addition to a group ID, the GFF format allows annotations to have a group class. This makes sure that all groups are unique even if they happen to share the same name. For example, you can have a GenBank accession named AP001234 and a clone named AP001234 and distinguish between them by giving the first one a class of Accession and the second a class of Clone.

You should use double-quotes around the group name or class if it contains white space.

A2. Creating a GFF table

The first 8 fields of the GFF format are easy to understand. The group field is a challenge. It is used in three distinct ways:

  1. to group together a single sequence feature that spans a discontinuous range, such as a gapped alignment.
  2. to name a feature, allowing it to be retrieved by name.
  3. to add one or more notes to the annotation.

1. Using the Group field for simple features

For a simple feature that spans a single continuous range, choose a name and class for the object and give it a line in the GFF file that refers to its start and stop positions.

 Chr3   giemsa heterochromatin  4500000 6000000 . . .   Band 3q12.1

2. Using the Group field to group features that belong together

For a group of features that belong together, such as the exons in a transcript, choose a name and class for the object. Give each segment a separate line in the GFF file but use the same name for each line. For example:

 IV     curated exon    5506900 5506996 . + .   Transcript B0273.1
 IV     curated exon    5506026 5506382 . + .   Transcript B0273.1
 IV     curated exon    5506558 5506660 . + .   Transcript B0273.1
 IV     curated exon    5506738 5506852 . + .   Transcript B0273.1

These four lines refer to a biological object of class "Transcript" and name B0273.1. Each of its parts uses the method "exon", source "curated". Once loaded, the user will be able to search the genome for this object by asking the browser to retrieve "Transcript:B0273.1". The browser can also be configured to allow the Transcript: prefix to be omitted.

You can extend the idiom for objects that have heterogeneous parts, such as a transcript that has 5' and 3' UTRs

 IV     curated  mRNA   5506800 5508917 . + .   Transcript B0273.1; Note "Zn-Finger"
 IV     curated  5'UTR  5506800 5508999 . + .   Transcript B0273.1
 IV     curated  exon   5506900 5506996 . + .   Transcript B0273.1
 IV     curated  exon   5506026 5506382 . + .   Transcript B0273.1
 IV     curated  exon   5506558 5506660 . + .   Transcript B0273.1
 IV     curated  exon   5506738 5506852 . + .   Transcript B0273.1
 IV     curated  3'UTR  5506852 5508917 . + .   Transcript B0273.1

In this example, there is a single feature with method "mRNA" that spans the entire range. It is grouped with subparts of type 5'UTR, 3'UTR and exon. They are all grouped together into a Transcript named B0273.1. Furthermore the mRNA feature has a note attached to it.

*NOTE* The subparts of a feature are in absolute (chromosomal or contig) coordinates. It is not currently possible to define a feature in absolute coordinates and then to load its subparts using coordinates that are relative to the start of the feature.

Some annotations do not need to be individually named. For example, it is probably not useful to assign a unique name to each ALU repeat in a vertebrate genome. For these, just leave the Group field empty.

3. Using the Group field to add a note

The group field can be used to add one or more notes to an annotation. To do this, place a semicolon after the group name and add a Note field:

 Chr3 giemsa heterochromatin 4500000 6000000 . . . Band 3q12.1 ; Note "Marfan's syndrome"

You can add multiple Notes. Just separate them by semicolons:

  Band 3q12.1 ; Note "Marfan's syndrome" ; Note "dystrophic dysplasia"

The Note should come AFTER the group type and name.

3. Using the Group field to add an alternative name

If you want the feature to be quickly searchable by an alternative name, you can add one or more Alias tags. A feature can have multiple aliases, and multiple features can share the same alias:

 Chr3 giemsa heterochromatin 4500000 6000000 . . . Band 3q12.1 ; Alias MFX

Searches for aliases will be both faster and more reliable than searches for keywords in notes, since the latter relies on whole-text search methods that vary somewhat from DBMS to DBMS.

A3. Identifying the reference sequence

Each reference sequence in the GFF table must itself have an entry. This is necessary so that the length of the reference sequence is known.

For example, if "Chr1" is used as a reference sequence, then the GFF file should have an entry for it similar to this one:

 Chr1 assembly chromosome 1 14972282 . + . Sequence Chr1

This indicates that the reference sequence named "Chr1" has length 14972282 bp, method "chromosome" and source "assembly". In addition, as indicated by the group field, Chr1 has class "Sequence" and name "Chr".

It is suggested that you use "Sequence" as the class name for all reference sequences, since this is the default class used by the Bio::DB::GFF module when no more specific class is requested. If you use a different class name, then be sure to indicate that fact with the "reference class" option (see below).

A4. Sequence alignments

There are several cases in which an annotation indicates the relationship between two sequences. One common one is a similarity hit, where the annotation indicates an alignment. A second common case is a map assembly, in which the annotation indicates that a portion of a larger sequence is built up from one or more smaller ones.

Both cases are indicated by using the Target tag in the group field. For example, a typical similarity hit will look like this:

 Chr1 BLASTX similarity 76953 77108 132 + 0 Target Protein:SW:ABL_DROME 493 544

Here, the group field contains the Target tag, followed by an identifier for the biological object. The GFF format uses the notation Class:Name for the biological object, and even though this is stylistically inconsistent, that's the way it's done. The object identifier is followed by two integers indicating the start and stop of the alignment on the target sequence.

Unlike the main start and stop columns, it is possible for the target start to be greater than the target end. The previous example indicates that the the section of Chr1 from 76,953 to 77,108 aligns to the protein SW:ABL_DROME starting at position 493 and extending to position 544.

A similar notation is used for sequence assembly information as shown in this example:

 Chr1        assembly Link   10922906 11177731 . . . Target Sequence:LINK_H06O01 1 254826
 LINK_H06O01 assembly Cosmid 32386    64122    . . . Target Sequence:F49B2       6 31742

This indicates that the region between bases 10922906 and 11177731 of Chr1 are composed of LINK_H06O01 from bp 1 to bp 254826. The region of LINK_H0601 between 32386 and 64122 is, in turn, composed of the bases 5 to 31742 of cosmid F49B2.

A6. Loading the GFF file into the database

Use the BioPerl script utilities bulk_load_gff.pl, load_gff.pl or (if you are brave) fast_load_gff.pl to load the GFF file into the database. For example, if your database is a MySQL database on the local host named "dicty", you can load it into an empty database using bulk_load_gff.pl like this:

  bulk_load_gff.pl -c -d dicty my_data.gff

To update existing databases, use either load_gff.pl or fast_load_gff.pl. The latter is somewhat experimental, so use with care.

A5. Aggregators

The Bio::DB::GFF module has a feature known as "aggregators". These are small software packages that recognize certain common feature types and convert them into complex biological objects. These aggregators make it possible to develop intelligent graphical representations of annotations, such as a gene that draws confirmed exons differently from predicted ones.

An aggregator typically creates a new composite feature with a different method than any of its components. For example, the standard "alignment" aggregator takes multiple alignments of method "similarity", groups them by their name, and returns a single feature of method "alignment".

The various aggregators are described in detail in the Bio::DB::GFF manual page. It is easy to write new aggregators, and also possible to define aggregators on the fly in the gbrowse configuration file. It is suggested that you use the sample GFF files from the yeast, drosophila and C. elegans projects to see what methods to use to achieve the desired results.

In addition to the standard aggregators that are distributed with BioPerl, GBrowse distributes several experimental and/or special-purpose aggregators:

match_gap

This aggregator is used for GFF3 style gapped alignments, in which there is a single feature of method 'match' with a 'Gap' attribute. This aggregator was contributed by Dmitri Bichko.

orf

This aggregator aggregates raw "ORF" features into "coding" features. It is basically identical to the "coding" aggregator, except that it looks for features of type "ORF" rather than "cds".

reftranscript

This aggregator was written to make the compound feature, "reftranscript" for use with Gbrowse editing software developed outside of the GMOD development group. It can be used to aggregate "reftranscripts" from "refexons", loaded as second copy features. These features, in contrast to "transcripts", are usually implemented as features which cannot be edited and serve as starting point references for annotations added using Gbrowse for feature visualization.

Adding features to the compound feature, "reftranscript", can be done by adding to the "part_names" call (i.e. "refCDS").

waba_alignment

This aggregator handles the type of alignments produced by Jim Kent's WABA program, and was written to be compatible with the C elegans GFF files. It aggregates the following feature types into an aggregate type of "waba_alignment":

   nucleotide_match:waba_weak
   nucleotide_match:waba_strong
   nucleotide_match:waba_coding
wormbase_gene

This aggregator was written to be compatible with the C elegans GFF2 files distributed by the Sanger Institute. It aggregates raw "CDS", "5'UTR", "3'UTR", "polyA" and "TSS" features into "transcript" features. For compatibility with the idiosyncrasies of the Sanger GFF format, it expects that the full range of the transcript is contained in a main feature of type "Sequence".

It is strongly recommended that for mirroring C. elegans annotations, you use the "processed_transcript" aggregator in conjunction with the GFF3 files found at:

 ftp://ftp.wormbase.org/pub/wormbase/genomes/elegans/genome_feature_tables/GFF3

IT IS NOT NECESSARY TO USE AGGREGATORS WITH THE CHADO, BIOSQL OR BIO::DB::SEQFEATURE::STORE (GFF3) DATABASES.

B. ADDING A NEW DATABASE TO THE BROWSER ^

Each data source has a corresponding configuration file in the directory gbrowse.conf. Once you've created and loaded a new database, you should make a copy of one of the existing configuration files and modify it to meet your needs. The name of the new configuration file must follow the form:

  sourcename.conf

where "sourcename" is a short word that describes the data source. You can use this name to select the data source when linking to the browser. Just construct a URL that uses "sourcename" as a virtual directory under cgi-bin/gbrowse:

  http://your.site.org/cgi-bin/gbrowse/sourcename/

(Note: If you don't add the slash at the end, gbrowse will automatically do it for you, since the terminal slash is needed to work around an apparent bug in MSIE's cookie handling.)

It is suggested that you use the same name as the database, although this isn't a requirement. (If no "source=" argument is given, gbrowse picks the first configuration file that occurs alphabetically; you can control this by placing numbers in front of the configuration file, as in "01.yeast.conf".)

The configuration file is divided into a number of sections, each one introduced by a [SECTION TITLE]. The [GENERAL] section contains settings that are applicable to the entire application. Other sections define tracks to display.

I suggest that you begin with one of the example configuration files provided with the distribution and modify it to suit your needs.

B1. The [GENERAL] Section

The [GENERAL] section consists of a series of name=value options. For example, the beginning of the yeast.conf sample configuration file looks like this:

 [GENERAL]
 description = S. cerevisiae (via SGD Nov 2001)
 db_adaptor  = Bio::DB::GFF
 db_args     = -adaptor dbi::mysql
               -dsn     dbi:mysql:database=yeast;host=localhost
 aggregators = transcript alignment
 user        =
 passwd      =

Each option is a single word or phrase, usually in lower case. This is followed by an equals sign and the value of the option. You can add whitespace around the equals sign in order to increase readability. If a value is very long, you can continue it on additional lines provided that you put a tab or other whitespace on the continuation lines. For example:

 description = S. cerevisiae annotations via SGD Nov 2001, and
             converted using the process_sgd.pl script

Any lines that begin with a pound sign (#) are considered comments and ignored.

During this discussion, you might want to follow along with one of the example configuration files.

The following [GENERAL] options are recognized:

B2. The [TRACK DEFAULTS] section

The track defaults section specifies default values for each track. The following common options are recognized:

             glyph
             height
             bgcolor
             fgcolor
             fontcolor
             font2color
             strand_arrow

These options control the default graphical settings for any annotation types that are not explicitly specified. See the section below on controlling the settings. Any of the options allowed in the [track] sections described below are allowed here.

B3. Track Sections

Any other [Section] in the configuration file is treated as a declaration of a track. The order of track sections will become the default order of tracks on the display (the user can change this later). Here is a typical track declaration from yeast.conf:

 [Genes]
 feature      = gene:sgd
 glyph        = generic
 bgcolor      = yellow
 forwardcolor = yellow
 reversecolor = turquoise
 strand_arrow = 1
 height       = 6
 description  = 1
 key          = Named gene

This track is named "Genes". You may use a short mnemonic if you prefer; this will make the URL shorter when the user bookmarks a view he or she likes. Track names can contain almost any character, including whitespace, but cannot contain the "-" or "+" signs because these are used to separate track names in the URL when bookmarking. [My Genes] is OK, but [My-Genes] is not.

As in the general configuration section, the track declaration contains multiple name=value option pairs.

Valid options are as follows:

1 feature

This relates the track to one or more feature types as they appear in the database. Recall that each feature has a method and source. This is represented in the form method:source. So, for example, a feature of type "gene:sgd" has the method "gene" and the source "sgd".

It is possible to omit the source. A feature of type "gene" will include all features whose methods are "gene", regardless of the source field. It is not possible to omit the method.

It is possible to have several feature types displayed on a single track. Simply provide the feature option with a space-delimited list of the features you want to include. For example:

    feature = gene:sgd stRNA:sgd

This will include features of type "gene:sgd" and "stRNA:sgd" in the same track and display them in a similar fashion.

2 remote feature

This relates the track to a remote feature track somewhere on the Internet. The value is a http: or ftp: URL, and may correspond to a static file of features in GFF format, gbrowse upload format, a CGI script, or a DAS source. When this option is active, the "feature" option and most of the glyph control options described below are ignored, but the "citation" and "key" options are honored.

Example:

 remote feature = http://www.wormbase.org/cgi-bin/das/wormbase?type=mRNA
3 glyph

This controls the glyph (graphical icon) that is used to represent the feature. The list of glyphs and glyph-specific options are listed in the section GLYPHS AND GLYPH OPTIONS. The "generic" glyph is the default.

4 bgcolor

This controls the background color of the glyph. The format of colors is explained in GLYPHS AND GLYPH OPTIONS.

5 fgcolor

This controls the foreground color (outline color) of the glyph. The format of colors is explained in GLYPHS AND GLYPH OPTIONS.

6 fontcolor

This controls the color of the primary font of text drawn in the glyph. This is the font used for the features labels drawn at the top of the glyph.

7 font2color

This controls the color of the secondary font of text drawn in the glyph. This is the font used for the longish feature descriptions drawn at the bottom of the glyphs.

8 height

This option sets the height of the glyph. It is expressed in pixels.

9 strand_arrow

This is a true or false value, where true is 1 and false is 0. If this option is set to true, then the glyph will indicate the strandedness of the feature, usually by drawing an arrow of some sort. Some glyphs are inherently stranded, or inherently non-stranded and simply ignore this option.

10 label

This is a true or false value, where true is 1 and false is 0. If the option is set to true, then the name of the feature (i.e. its group name) is printed above the feature, space allowing.

11 description

This is a true or false value, where true is 1 and false is 0. If the option is set to true, then the description of the feature (any Note fields) is printed below the feature, space allowing.

12 key

This option controls the descriptive key that is drawn in the key area at the bottom of the image. It also appears in the checkboxes that the end user uses to switch tracks on and off. If not specified, it defaults to the track name.

13 citation

If present, this option creates a human-readable descriptive paragraph describing the feature and how it was derived. This is the text information that is displayed when the user clicks on the track name in the checkbox group. The value can either be a URL, in which case clicking on the track name invokes the corresponding URL, or a text paragraph, in which case clicking on the track name generates a page containing the text description. Long paragraphs can be continued across multiple lines, provided that continuation lines begin with whitespace.

14 link, title, link_target

These options are identical to the similarly-named options in the [GENERAL] section, but change the rules on a track-by-track basis. They can be used to override the global rules. To force a track not to contain any links, use a blank value.

15 box_subparts

If this option is greater than zero, then gbrowse will generate imagemap rectangles for each of the subparts of a feature (e.g. the exons within a transcript), allowing you to link each subpart separately. The numeric value will control the number of levels of subfeatures that the boxes will descend into. For example, if using the "gene" glyph, set -box_subparts to 2 to create boxes for the whole gene (level 0), the mRNAs (level 1) and the exons (level 2).

16 feature_low

If this option is present, GBrowse will use the list of feature types listed here at resolution views. (This is one of the ways that semantic zooming is implemented.) This allows you, for example, to switch off detailed exon, UTR, promoters and other within-the-gene features, and just show the start and stop of the transcription unit.

17 global feature

If this option is present and set to a true value (e.g. "1"), GBrowse will automatically generate a pseudo-feature that starts at the beginning of the currently displayed region and extends to the end. This is often used in conjunction with the "translation" and "dna" glyphs in order to display global characteristics of the sequence. If this option is set, then you do not need to specify a "feature" option.

18 group_pattern

This option lets you connect related features by dotted lines based on a pattern match in the features' names. A typical example is connecting the 5' and 3' read pairs from ESTs or plasmids. See GROUPING FEATURES for details.

19 group_on

For Bio::DB::SeqFeature::Store databases only, the group_on field allows you to group features together by display_name, target or any other method. This is mostly useful for XY-plot data, where you may want to dynamically group related data points together so that they share the same vertical scaling.

Example:

        group_on = display_name

(this feature is under refinement and may change in the future)

20 restrict

This option allows you to restrict who is allowed to view the current track by host name, IP address or username/password. See AUTHENTICATION AND AUTHORIZATION for details.

21 category

This option allows you to group tracks into different groups on the GBrowse display in addition to the default group called 'General'. For example, if you wanted several tracks to be in a separate group called "Genes", you would add this to each of the track defintions:

  category = Genes

Note that it is not possible to make subcategories. If all tracks are categorized, then the "General" category will not be displayed.

22 das category, das landmark, das subparts, das superparts

All these options pertain to exporting the GBrowse database as a DAS data source. Please see DAS_HOWTO for more information.

A large number of glyph-specific options are also recognized. These are described in the next section.

B4. Glyphs and Glyph Options

A large variety of glyphs are available, and more are being added as the Bio::Graphics module grows.

A list of the common glyphs and their options is provided by the GBrowse itself. Click on the "[Help]" link in the section labeled "Upload your own annotations". This page also lists the valid foreground and background colors. Most of the glyphs are found in the BioPerl distribution, but a few are distributed directly with GBrowse.

The most popular glyph types are:

  Glyph                 Description
  -----                 -----------

  generic               a rectangle
  allele_tower          allele found at a SNP position
  arrow                 an arrow
  anchored_arrow        a span with vertical bases |---------|.  If one
                        or the other end of the feature is off-screen, the
                        base will be replaced by an arrow.
  box                   another rectangle; doesn't show subparts of features
  cds                   shows the reading frame of spliced transcripts; used
                        in conjunction with the "coding" aggregator.
  diamond               a point-like feature represented as a triangle
  dna                   DNA and GC content
  heterogeneous_segments a multi-segmented feature in which each segment can
                        have a distinctive color.  For Jim Kent's WABA features,
                        this works with the waba_alignment aggregator.
  idiogram              this takes specially-formatted feature data and turns it
                        into an idiogram of a Giemsa-stained metaphase chromosome
  image                 this embeds photographic images and/or diagrams on features
  processed_transcript  multi-purpose representation of a spliced mRNA, including
                        positions of UTRs
  segments              a multi-segmented feature such as an alignment
  span                  like anchored_arrow, except that the ends are
                        truncated at the edge of the panel, not turned
                        into an arrow
  trace                 reads an SCF trace file and draws a graphic representation
  triangle              a point-like feature represented as a diamond
  transcript            a gene model
  transcript2           a slightly different representation of a gene model
  translation           1-, 3- and 6-frame translations
  wormbase_transcript   yet another gene model that can show UTR segments
                        (for features that conform to the WormBase gene
                        schema). Used in conjunction with the
                        "wormbase_gene" aggregator.
  xyplot                histograms and line plots

A more definitive list of glyph options can be found in the Bio::Graphics manual pages. Consult the manual pages for the following modules:

  Glyph                         Manual Page
  -----                         -----------

  (common options for all)      Bio::Graphics::Glyph
  allele_tower                  Bio::Graphics::Glyph::allele_tower
  anchored_arrow                Bio::Graphics::Glyph::anchored_arrow
  arrow                         Bio::Graphics::Glyph::arrow
  box                           Bio::Graphics::Glyph::box
  cds                           Bio::Graphics::Glyph::cds
  crossbox                      Bio::Graphics::Glyph::crossbox
  diamond                       Bio::Graphics::Glyph::diamond
  dna                           Bio::Graphics::Glyph::dna
  dot                           Bio::Graphics::Glyph::dot
  ellipse                       Bio::Graphics::Glyph::ellipse
  extending_arrow               Bio::Graphics::Glyph::extending_arrow
  generic                       Bio::Graphics::Glyph::generic
  graded_segments               Bio::Graphics::Glyph::graded_segments
  heterogeneous_segments        Bio::Graphics::Glyph::heterogeneous_segments
  idiogram                      Bio::Graphics::Glyph::idiogram
  image                         Bio::Graphics::Glyph::image
  line                          Bio::Graphics::Glyph::line
  primers                       Bio::Graphics::Glyph::primers
  processed_transcript          Bio::Graphics::Glyph::processed_transcript
  rndrect                       Bio::Graphics::Glyph::rndrect
  ruler_arrow                   Bio::Graphics::Glyph::ruler_arrow
  segments                      Bio::Graphics::Glyph::segments
  span                          Bio::Graphics::Glyph::span
  toomany                       Bio::Graphics::Glyph::toomany
  trace                         Bio::Graphics::Glyph::trace
  transcript                    Bio::Graphics::Glyph::transcript
  transcript2                   Bio::Graphics::Glyph::transcript2
  translation                   Bio::Graphics::Glyph::translation
  triangle                      Bio::Graphics::Glyph::triangle
  wormbase_transcript           Bio::Graphics::Glyph::wormbase_transcript
  xyplot                        Bio::Graphics::Glyph::xyplot

The "perldoc" command is handy for reading the documentation from the Unix command line. For example:

   perldoc Bio::Graphics::Glyph::primers

This will provide you with a summary of the options that apply to the "primers" glyph.

In the manual pages, the glyph options are presented the way they are called from Perl. For example, the documentation will tell you to use the -connect_color option to set the color to use when drawing the line that connects the two inward pointing arrows in the primer pair glyph. This translates to the configuration file as an option named "connect_color". For example:

 [PCR Products]
 glyph = primer
 connect_color = blue

When referring to colors, you can use a variety of color names such as "blue" and "green". To get the full list, cut and paste the following magic incantation into the command line:

 perl -MBio::Graphics::Panel -e 'print join "\n",Bio::Graphics::Panel->color_names'

or see this URL:

  http://www.wormbase.org/db/seq/gbrowse?help=annotation

Alternatively, you can use the #RRGGBB notation to specify the red, green and blue components of the color. Refer to any book on HTML for the details on using the notation.

B5. Adding features to the overview

You can make any set of tracks appear in the overview by creating a stanza with a title of the format [<label>:overview], where <label> is any unique label of your choice. The format of the stanza is identical to the others, but the indicated track will appear in the overview rather than as an option in the detailed view. For example, this stanza adds to the overview a set of features of method "gene", source "framework":

 [framework:overview]
 feature       = gene:framework
 label         = 1
 glyph         = generic
 bgcolor       = lavender
 height        = 5
 key           = Mapped Genes

Similarly, you can make a track appear in the region panel by appending ":region" to its name:

 [genedensity:region]
 feature       = gene_density
 glyph         = xyplot
 graph_type    = boxes
 scale         = right
 bgcolor       = red
 fgcolor       = red
 height        = 20
 key           = SNP Density

B6. Semantic Zooming

Sometimes you will want to change the appearance of a track when the user has zoomed out or zoomed in beyond a certain level. To indicate this, create a set of "length qualified" stanzas of format [<label>:<zoom level>], where all stanzas share the same <label>, and <zoom level> indicates the minimum size of the region that the stanza will apply to. For example:

  [gene]
  feature = transcript:curated
  glyph    = dna
  fgcolor  = blue
  key      = genes
  citation = example semantic zoom track

  [gene:500]
  feature = transcript:curated
  glyph   = transcript2

  [gene:100000]
  feature = transcript:curated
  glyph   = arrow

  [gene:500000]
  feature = transcript:curated
  glyph   = generic

This series of stanzas says to use the "transcript2" glyph when the segment being displayed is 500 bp or longer, to use the "arrow" glyph when the segment being displayed is 100,000 bp or longer, and the "generic" glyph when the region being displayed is 500,000 bp or longer. For all other segment lengths (1 to 499 bp), the ordinary [gene] stanza will be consulted, and the "dna" glyph will be displayed. The bare [gene] stanza is used to set all but the "feature" options for the other stanzas. This means that the fgcolor, key and citation options are shared amongst all the [gene:XXXX] stanzas, but the "feature" option must be repeated.

You can override any options in the length qualified stanzas. For example, if you want to change the color to red in when displaying genes on segments between 500 and 99,999 bp, you can modify the [gene:500] stanza as follows:

  [gene:500]
  feature = transcript:curated
  glyph   = transcript2
  fgcolor = red

It is also possible to display different features at different zoom levels, although you should handle this potentially confusing feature with care.

If you wish to turn off a track entirely, you can use the "hide" flag to hide the track when the display exceeds a certain size:

  [6_frame_translation:50000]
  hide = 1

B7. Computed Options

Some options can be computed at run time by using Perl subroutines as their values. These are known as "callbacks." Currently this works with the values of the "link", "title", "link_target", "header" and "footer" options, and any glyph-specific option that appears in a track section.

You need to know the Perl programming language to take advantage of this. The general format of this type of option is:

  option name = sub {
              some perl code;
              some more perl code;
              even more perl code;
              }

The value must begin with the sequence "sub {" in order to be recognized as a subroutine declaration. After this, you can have one or more lines of Perl code followed by a closing brace. Continuation lines must begin with whitespace.

When the browser first encounters an option like this one, it will attempt to compile it into Perl runtime code. If successful, the compiled code will be stored for later use and invoked whenever the value of the option is needed. (Otherwise, an error message will appear in your server error log).

For options of type "footer" and "header", the subroutine is passed no arguments. It is expected to produce some HTML and return it as a string value.

For glyph-specific features, such as "bgcolor" the subroutine will be called at run time with five arguments consisting of the feature, the name of the option, the current part number of the feature, the total number of parts in this feature, and the glyph corresponding to the feature. Usually you will just look at the first argument. The return value is treated as the value of the corresponding option. For example, this bgcolor subroutine will call the feature's primary_tag() method, and return "blue" if it is an exon, "orange" otherwise:

  bgcolor = sub {
          my $feature = shift;
          return "blue" if $feature->primary_tag eq 'exon';
          return "orange";
          }

See the manual page for Bio::DB::GFF::Feature for information on how to interrogate the feature object.

For special effects, such as coloring the first and last exons differently, you may need access to all five arguments. Here is an example that draws the first and last parts of a feature in blue and the rest in red:

   sub { 
         my($feature,$option_name,$part_no,$total_parts,$glyph) = @_;
         return 'blue' if $part_no == 0;                # zero-based indexing!
         return 'blue' if $part_no == $total_parts-1;   # zero-based indexing!
         return 'red';
         }

See the Bio::Graphics::Panel manual page for more details.

Callbacks for the "link", "title", and "link_target" options have a slightly different call signature. They receive three arguments consisting of the feature, the Bio::Graphics::Panel object, and the Bio::Graphics::Glyph object corresponding to the current track within the panel:

  link = sub {
             my ($feature, $panel, $track) = @_;
             ... do something
             }

Ordinarily you will only need to use the feature object. The other arguments are useful to look up panel-specific settings such as the pixel width of the panel or the state of the "flip" setting:

  title = sub {
          my ($feature,$panel,$track) = @_;
          my $name = $feature->display_name;
          return $panel->flip ? "$name (flipped)" : $name;
       }

Named Subroutine References ---------------------------

If you use a version of BioPerl after April 15, 2003, you can also use references to named subroutines as option arguments. To use named subroutines, add an init_code section to the [GENERAL] section of the configuration file. init_code should contain nothing but subroutine definitions and other initialization routines. For example:

  init_code = sub score_color {
                my $feature = shift;
                if ($feature->score > 50) { 
                  return 'red';
                } else {
                  return 'green';
                }
              }
              sub score_height {
                my $feature = shift;
                if ($feature->score > 50) { 
                  return 10;
                } else {
                  return 5;
                }
              }

Then simply refer to these subroutines using the \&name syntax:

    [EST_ALIGNMENTS]
    glyph = generic
    bgcolor = \&score_color
    height  = \&score_height

You can declare global variables in the init_code subroutine if you use "no strict 'vars';" at the top of the section:

    init_code = no strict 'vars';
                $HEIGHT = 10;
                sub score_height {
                  my $feature = shift;
                  $HEIGHT++;
                  if ($feature->score > 50) { 
                    return $HEIGHT*2;
                  } else {
                    return $HEIGHT;
                  }
                }

Due to the way the configuration file is parsed, there must be no empty lines in the init_code section. Either use comments to introduce white space, or "use" a .pm file to do anything fancy.

Subroutines that you define in the init_code section, as well as anonymous subroutines, will go into a package that changes unpredictably each time you load the page. If you need a predictable package name, you can define it this way:

   init_code = package My; sub score_height { .... }

   [EST_ALIGNMENTS]
   height = \&My::score_height

B8. Declaring New Aggregators

The Bio::DB::GFF data model recognizes a single-level of "grouping" of features, but doesn't specify how to use the group information to correctly assemble the various individual components into a biological object. Aggregators are used to assemble this information. For example, let's say that you decide that your preferred "transcript" data model contains three subfeature types: a set of one or more features of method "exon", a single feature of method "TSS", and a single feature of method "polyA". Optionally, the data model could contain a single "main subfeature" that runs the length of the entire transcript. We might give this feature a method of "primary_transc" (for "primary transcript.")

In a GFF file, a three-exon transcript might be represented as follows:

 Chr1 confirmed primary_transc 100 500  .  +  .  Transcript "ABC.1"
 Chr1 confirmed TSS            100 100  .  +  .  Transcript "ABC.1"
 Chr1 confirmed exon           100 200  .  +  .  Transcript "ABC.1"
 Chr1 confirmed exon           250 300  .  +  .  Transcript "ABC.1"
 Chr1 confirmed exon           400 500  .  +  .  Transcript "ABC.1"
 Chr1 confirmed polyA          500 500  .  +  .  Transcript "ABC.1"

To aggregate this, you would like to create an aggregator named "transcript", whose "main method" is "primary_transc", and whose "sub methods" are "TSS," "exon," and "polyA."

The way to indicate this in the configuration file is to add a "complex aggregator" to the list of aggregators:

  aggregator = transcript{TSS,exon,polyA/primary_transc}

The format of this value is "aggregator_name{submethod1,submethod2,.../mainmethod}".

You can now use the name of the aggregator name as the argument of the "feature" option in a track section:

  [Transcripts]
  feature      = transcript
  glyph        = segments
  bgcolor      = wheat
  fgcolor      = black
  height       = 10
  key          = Transcripts

If you do not have a main subfeature, leave off the "/mainmethod". For example:

  aggregator = transcript{TSS,exon,polyA}

A few formatting notes. You are free to mix simple and complex aggregators in the "aggregator" option. For example, you can activate the standard "clone" and "alignment" aggregators as well as the new transcript aggregator with a line like this one:

 aggregator = clone
              transcript{TSS,exon,polyA/primary_transc}
              alignment

If the complex aggregator contains whitespace or apostrophes, you must surround it with double-quotes, like this:

   "transcript{TSS,5'UTR,3'UTR,exon,polyA/primary_transc}"

Be aware that some glyphs look for particular method names when rendering aggregated features. For example, the standard "transcript" glyph is closely tied to the "transcript" aggregator, and looks for submethods named "intron", "exon" and "CDS", and a main method named "transcript."

Here is the list of available predefined aggregators:

     alignment
     clone
     coding
     transcript
     none
     orf
     waba_alignment
     wormbase_gene

To view the documentation for any of these aggregators, run the command "perldoc Bio::DB::GFF::Aggregator::aggregator_name", where "aggregator_name" is the name of the aggregator.

B9. GROUPING FEATURES

gbrowse recognizes the concept of a "group" of related features that are connected by dotted lines. The canonical example is a pair of ESTs that are related by being from the two ends of the same cDNA clone. However many feature databases, including the GFF database recommended for gbrowse, do not allow for arbitrary hierarchical grouping. To work around this, you may specify a feature name-based regular expression that will be used to trigger grouping.

It works like this. Say you are working with EST feature pairs and they follow the nomenclature 501283.5 and 501283.3, where the suffix is "5" or "3" depending on whether the read was from the 5' or 3' ends of the insert. To group these pairs by a dotted line, specify the "group_pattern" option in the appropriate track section:

      group_pattern =  /\.[53]$/

At render time, gbrowse will strip off this pattern from the names of all features in the EST track and group those that have a common base name. Hence 501283.5 and 501283.3 will be grouped together by a dotted line, because after the pattern is removed, they will share the same common name "501283".

This works for all embedded pattern, provided that stripping out the pattern results in related features sharing the same name. For example, if the convention were "est.for.501283" and "est.rev.501283", then this grouping pattern would have the desired effect:

      group_pattern = /\.(for|rev)\./

Don't forget to escape regular expression meta-characters and to consider the various ways in which the regular expression might break. It is entirely possible to create an invalid regular expression, in which case gbrowse will crash until you comment out the offending option.

B10. Controlling the gbrowse_details page

If a track definition's "link" option (see section B2) is set to AUTO, the gbrowse_details script will be invoked when the user clicks on a feature contained within the track. This will generate a simple table of all feature information available in the database. This includes the user-defined tag/value attributes set in Column 9 of the GFF for that feature.

You can control, to some extent, the formatting of the tag value table by providing a configuration stanza with the following format:

  [feature_type:details]
  tag1 = formatting rule
  tag2 = formatting rule
  tag3 = formatting rule

"feature_type" is the type of the feature you wish to control. For example, "gene:sgd" or simply "gene". You may also specify a feature_type of "default" to control the formatting for all features. "tag1", "tag2" and so forth are the tags that you wish to control the formatting of. The tags "Name," "Class", "Type", "Source", "Position", and "Length" are valid for all features, while "Target" and "Matches" are valid for all features that have a target alignment. In addition, you can use the names of any attributes that you have defined. Tags names are NOT case sensitive, and you may use a tag named "default" to define a formatting rule that is general to all tags (more specific formatting rules will override less specific ones).

A formatting rule can be a string with (possible) substitution values, or a callback. If a string, it can contain one or more of the substitution variable "$name", "$start", "$end", "$stop", "$strand", "$method", "$type", "$description" and "$class", which are replaced with the corresponding values from the current feature. In addition, the substitution variable "$value" is replaced with the current value of the attribute, and the variable "$tag" is replaced with the current tag (attribute) name. HTML characters are passed through.

For example, here is a simple way to boldface the Type field, italicize the Length field, and turn the Notes into a Google search:

 [gene:details]
 Type   = <b>$value</b>
 Length = <i>$value</b>
 Note  = <a href="http://www.google.com/search?q=$value">$value</a>

If you provide a callback, the callback subroutine will be invoked with three arguments. WARNING: the three arguments are different from the ones passed to other callbacks, and consist of the tag value, the tag name, and the current feature:

  Note = sub {
             my($value,$tag_name,$feature) = @_;
             do something....
             }

You can use this feature to format sequence attributes nicely. For example, if your features have a Translation attribute which contains their protein translations, then you are probably unsatisified with the default formatting of these features. You can modify this with a callback that word-wraps the value into lines of at most 60 characters, and puts the whole thing in a <pre> section.

 [gene:details]
 Translation = sub {
                my $value = shift;
                $value =~ s/(\S{1,60})/$1\n/g;
                "<pre>$value</pre>";
             }

B11. Linking out from gbrowse_details

The formatting rule mechanism described in the previous section is the recommended way of creating a link out from the gbrowse_details page. However, an older mechanism is available for backward compatibility.

To use this legacy mechanism, create a stanza header named [TagName:DETAILS], where TagName is the name of the tag (attribute name) whose values you wish to turn into URLs, and where DETAILS must be spelled with capital letters. Put the option "URL" inside this stanza, containing a string to be transformed into the URL.

For example, to link to a local cgi script from the following GFF line:

 IV     curated exon    518     550     . + .   Transcript B0273.1; local_id 11723

one might add the following stanza to the configuration file:

    [local_id:DETAILS]
    URL   = http://localhost/cgi-bin/localLookup.cgi?tag=$tag;id=$value

The URL option's value should be a URL containing one or more variables. Variables begin with a dollar sign ($), and are replaced at run time with the information relating to the selected feature attribute. Recognized variables are:

     $tag        The "tag" of the tag/value pair
     $value      The "value" of the tag/value pair

The value of URL can also be an anonymous subroutine, in which case the subroutine will be invoked with a two-element argument list consisting of the name of the tag and its value. This example, provided by Cyril Pommier, will convert Dbxref tags into links to NCBI, provided that the value of the tag looks like an NCBI GI number:

 [Dbxref:DETAILS]
 URL = sub { 
       my ($tag,$value)=@_;
       if ($value =~ /NCBI_gi:(.+)/){
        return "http://www.ncbi.nlm.nih.gov/gquery/gquery.fcgi?term=$1";
        }
        return;
      }

C. GENERATING HISTOGRAMS ^

With a little bit of additional effort, you can set one or more tracks up to display a density histogram of the features contained within the track. For example, the human data source in GBrowse demo (http://www.wormbase.org/db/seq/gbrowse/human) uses density histograms in the chromosomal overview. In addition, when the features in the SNP track become too dense to view, this track converts into a histogram. To see this in action, turn on the SNP track and then zoom out beyond 150K.

There are four steps for making histograms:

  1. generate the density data using the bp_generate_histogram.pl script.
  2. load the density data using load_gff.pl or fast_load_gff.pl.
  3. declare a density aggregator to the gbrowse configuration file
  4. add the density aggregator to the appropriate track in the configuration file.

The first step is to generate the density data. Currently this is done by generating a GFF file containing a set of "bin" feature types. Use the bp_generate_histogram.pl script to do this. You will find it in bioperl under the scripts/Bio-DB-GFF directory.

Assuming that your database is named "dicty", you have a feature named SNP, and you wish to generate a density distribution across 10,000 bp bins, here is the command you would use:

  bp_generate_histogram.pl -merge -d dicty -bin 10000 SNP >snp_density.gff

This is saying to use the "dicty" database (-d) option, to use 10,000 bp bins (the -bin option) and to count the occurrences of the SNP feature throughout the database. In addition, the -merge option says to merge all types of SNPs into a single bin. Otherwise they will be stratified by their source. The resulting GFF file contains a series of entries like these ones:

  Chr1  SNP bin 1     10000 49 + . bin Chr1:SNP
  Chr1  SNP bin 10001 20000 29 + . bin Chr1:SNP

What this is saying is that there are now a series of pseudo-features of type "bin:SNP" that occupy successive 10,000 bp regions of the genome. The score field contains the number of times a SNP was seen in that bin.

You'll now load this file using load_gff.pl or fast_load_gff.pl:

  load_gff.pl -d dicty snp_density.gff

The next step is to tell GBrowse how to use this information. You do this by creating a new aggregator for the SNP density information. Open the GBrowse configuration file and find the aggregators option. Add a new aggregator that looks like this:

  aggregators = snp_density{bin:SNP}

This is declaring a new feature named "snp_density" that is composed of subparts of type bin:SNP.

The last step is to declare a track for the density information. You will use the "xyplot" glyph, which can draw a number of graphs, including histograms. To add the SNP density information as a static track in the overview, create a section like this one:

 [SNP:overview]
 feature       = snp_density
 glyph         = xyplot
 graph_type    = boxes
 scale         = right
 bgcolor       = red
 fgcolor       = red
 height        = 20
 key           = SNP Density

This is declaring a new constant track in the overview named "SNP Density." The feature is "snp_density", corresponding to the aggregator declared earlier. The glyph is "xyplot" using the graph type of "boxes" to generate a column graph.

To set up a track so that the histogram appears when the user zooms out beyond 100,000 bp but shows the detailed information at higher magnifications, generate two track sections like these:

  [SNPs]
  feature       = snp
  glyph         = triangle
  point         = 1
  orient        = N
  height        = 6
  bgcolor       = blue
  fgcolor       = blue
  key           = SNPs

  [SNPs:100000]
  feature       = snp_density
  glyph         = xyplot
  graph_type    = boxes
  scale         = right

The first track section sets up the defaults for the SNP track. SNPs are represented as blue triangles pointing North. The second track declaration declares that when the user zooms out to over 100K base pairs, GBrowse should display the snp_density feature using the xyplot glyph.

D. INTERNATIONALIZATION ^

GBrowse is partially internationalized. End-users whose browsers are set to request a non-English language will see the GBrowse main and secondary screens in their preferred language, provided that GBrowse has the appropriate translation file.

Translation files are located in gbrowse.conf/languages/ and use the standard two-letter language abbreviations, such as "fr" for French, as well as the regional abbregiations, such as fr-CA for Canadian French. Currently there are translation files for French, Italian, and Japanese. If your favorite language isn't supported, you are encouraged to create a new translation file and contribute it to the GBrowse development effort. Please contact Lincoln Stein (lstein@cshl.org) for help in doing this.

If the end user does not specify a preferred language, GBrowse will default to "en" (English). You can change this by placing a "language" option in the configuration file somewhere inside the [GENERAL] section. For example, to make Japanese the default, create this entry:

  language = ja

GBrowse will still use the end-user's preferred language in preference to the default if the preferred language is available.

Although GBrowse automatically changes the text and button language, it can't automatically translate the track labels. If you would like the track labels to localize, you will have to provide your own translations in the "key", "citation" and "category" options. The syntax is similar to that used for semantic zooming:

  [gene]
  glyph   = transcript
  feature = transcript:curated
  height  = 10
  key     = Named Gene
  key:fr  = Gènes Nommés
  key:it  = I Geni dati un nome a
  key:sp  = Los Genes denominados
  category = Genes
  category:fr = Gènes

The option is followed by a colon and the two-letter language name to indicate that when the page is being displayed with this language, to use the indicated value of the option. The option without the colon is the default. You may enter accented and umlauted characters directly, as shown, or use the HTML entities. Non-English character sets, such as Japanese, should also work correctly, provided that the translation file indicates the correct character set to use.

HELP FILES:

The GBrowse help files are in English. Although there is support for internationalizing the hep files, no one has done this yet. If you are industrious and wish to translate the help files into your favorite language, find the two help files where they are located in htdocs/gbrowse/. One is named general_help.html, while the other is named annotation_help.html. Translate them, and create new files with the language prefix appended to the end. For example, the French translation of annotation_help.html would be annotation_help.html.fr.

LIMITATIONS:

- There is no localization support. For example, GBrowse will print large numbers using commas (e.g. 1,234,567) instead of periods, even when talking to a European browser.

- Although the HTML frame around the GBrowse genome image will use the appropriate character set, the overview and detail images themselves are limited to Latin alphabets. This is because of limited native character support in the GD library used by GBrowse. When a non-Latin character set is called for, such as Japanese, GBrowse will use Japanese for the frame, but English for the image.

- The rate at which the GBrowse team adds new features to the browser often outstrips the ability of volunteers to update the translation files. This means that new buttons and fields may be displayed in English on an otherwise correctly internationalized page.

E. AUTHENTICATION AND AUTHORIZATION ^

You can restrict who has access to gbrowse by IP address, host name, domain or username and password. Restriction can apply to the database as a whole, or to particular annotation tracks.

To limit access to a whole database, you can use Apache's standard authentication and authorization. Gbrowse uses a URL of this form to select which database it is set to:

      http://your.host/cgi-bin/gbrowse/your_database

where "your_database" is the name of the currently selected database. For example, the yeast database is http://your.host/cgi-bin/gbrowse/yeast.

To control access to the entire database, create a <Location> section in httpd.conf. The <Location> section should look like this:

   <Location /cgi-bin/gbrowse/your_database>
        Order deny,allow
        deny from all
        allow from localhost .cshl.edu .ebi.ac.uk
   </Location>

This denies access to everybody except for "localhost" and browsers from the domains .cshl.edu and .ebi.ac.uk. You can also limit by IP address, by username and password or by combinations of these techniques. See http://httpd.apache.org/docs/howto/auth.html for the full details.

You can also limit individual tracks to certain individuals or organizations. Unless the stated requirements are met, the track will not appear on the main screen or any of the configuration screens. To set this up, add a "restrict" option to the track you wish to make off-limits:

        [PROPRIETARY]
        feature = etc
        glyph   = etc
        restrict = Order deny,allow
                   deny from all
                   allow from localhost .cshl.edu .ebi.ac.uk

The value of the restrict option is identical to the Apache authorization directives and can include any of the directives "Order," "Satisfy," "deny from," "allow from," "require valid-user" or "require user." The only difference is that the "require group" directive is not supported, since the location of Apache's group file is not passed to CGI scripts. Note that username/password authentication must be turned on in httpd.conf and the user must have successfully authenticated himself in order for the username to be available.

As with other gbrowse options, restrict can be a code subroutine. The subroutine will be called with three arguments consisting of the host, ip address and authenticated user. It should return a true value to allow access to the track, or a false value to forbid it. This can be used to implement group-based authorization or more complex schemes.

Here is an example that uses the Text::GenderFromName to allow access if the user's name sounds female and forbids access if the name sounds male. (It might be useful for an X-chromosome annotation site.)

    restrict = sub {
               my ($host,$ip,$user) = @_;
               return unless defined $user;
               use Text::GenderFromName qw(gender);
               return gender($user) eq 'f';
             }

You should be aware that the username will only be defined if username authentication is turned on and the user has successfully authenticated himself against Apache's user database using the correct password. In addition, the hostname will only be defined if HostnameLookups have been turned on in httpd.conf. In the latter case, you can convert the IP address into a hostname using this piece of code:

    use Socket;
    $host = gethostbyaddr(inet_aton($addr),AF_INET);

Note that this may slow down the response time of gbrowse noticeably if you have a slow DNS name server.

Another thing to be aware of when restricting access to an entire database is that that even though the database itself will not be accessible to unauthorized users, the name of the database will still be available from the popup "Data Source" menu. If you wish even the name to be suppressed from view by unauthorized users, add the following line to the [GENERAL] section of the configuration file of the database you wish to suppress:

    restrict = require valid-user

The syntax described earlier for restricting access to tracks by hostname, IP address or username holds true for restricting the visibility of the database on the Data Source popup menu.

F. DISPLAYING GENETIC AND RH MAPS ^

GBrowse can be tweaked to make it more suitable for displaying genetic and radiation hybrid maps.

The main issue is that the Bio::DB::GFF database expects coordinates to be positive integers, not fractions, but genetic and RH maps use floating point numbers. Working around this is a bit of an ugly hack. Before loading your data you must multiply all your coordinates by a constant power of 10 in order to convert them into integers. For example, if a genetic map uses Morgan units ranging from 0 to 1.80, you would multiple by 100 to create a map in ranging from 0 to 180.

Create a GFF file containing the markers in modified coordinates and load it as usual. Now you must tell GBrowse to reverse these changes. Enter the following options into the [GENERAL] section of the configuration file:

 units = M
 unit_divider = 100

These two options tell GBrowse to use "M" (Morgan) units, and to divide all coordinates by 100. GBrowse will automatically display the scale using the most appropriate units, so the displayed map will typically be drawn using cM units.

G. CHANGING THE LOCATION OF THE CONFIGURATION FILES ^

If you wish to change the location of the gbrowse.conf configuration file directory, you must manually edit the gbrowse CGI script. Open the script in a text editor, and find this section:

 ###################################################################
 # Non-modperl users should change this variable if needed to point
 # to the directory in which the configuration files are stored.
 #
 use constant CONF_DIR => '/usr/local/apache/conf/gbrowse.conf';
 #
 ###################################################################

Change the definition of CONF_DIR to the desired location of the configuration files.

An alternative, for users of mod_perl only, is to add the GBrowseConf per-directory variable to the configuration for the directory in which the gbrowse script lives. This variable overrides the CONF_DIR value. For example:

 <Directory /usr/local/apache/cgi-perl>
   SetHandler      perl-script
   PerlHandler     Apache::Registry
   PerlSendHeader  On
   Options         +ExecCGI
   PerlSetVar      GBrowseConf /etc/gbrowse.conf
 </Directory>

H. USING DAS (DISTRIBUTED ANNOTATION SYSTEM) DATABASES ^

You may insert features from a DAS source into any named track. Create a stanza as usual but instead of specifying the feature type using the "feature" option, give the desired DAS URL using the "remote feature" option:

 remote feature = http://dev.hapmap.org/cgi-perl/das/t2d_testing?type=ldblock

Because DAS sources specify the glyph and visualization options, most of the settings such as bgcolor will be ignored. However, the track key and citation options are honored.

You can use the same syntax to load a GFF file or a feature file in Gbrowse upload format into a track. Just provide a URL that returns the desired data.

You can also run GBrowse entirely off a single DAS source. To get this support, you must use Bio::Das version 0.90 or higher, available from http://www.biodas.org.

A sample [GENERAL] configuration section looks like this:

 [GENERAL]
 description   = Das Example Database (dicty)
 db_adaptor    = Bio::Das
 db_args       = -source http://www.biodas.org/cgi-bin/das 
                 -dsn    dicty

The db_adaptor option must be set to "Bio::Das". The db_args option must contain a -source pointing to the base of the remote DAS server, and a -dsn pointing to the name of the annotation database.

The remainder of the configuration file should be configured as described earlier. The following short script will return a list of the feature types known to the remote DAS server. You can use the output of this script as the basis for the tracks to configure.

 #!/usr/bin/perl

 use strict;

 use Bio::Das;
 my $db = Bio::Das->new('http://localhost/cgi-bin/das'=>'dicty');
 print join "\n",$db->types;

Limitations:

The DAS implementation does not descend into subcomponents. For example, if the user requests features on a chromosome, but the remote DAS server has annotated genes using contig coordinates, then the genes will not appear on the chromosome.

The gbrowse_details script does not provide useful information because the DAS/1 protocol does not provide a way to retrieve attribute information on a named feature.

I. THE BioMOBY BROWSER ^

The BioMOBY project aims to design and deploy platforms that enable and simplify biological database interoperability.

To date, the MOBY-Services (MOBY-S) branch of the BioMOBY project has published a fairly stable API that is now being used by data providers worldwide to publish their data in an interoperable manner. A simple MOBY browser has been written for Gbrowse that allows the end-user to "surf" out of their Gbrowse view and begin exploring data related to the genomic features displayed in Gbrowse.

Configuration of the gbrowse_moby script does, at this time, require some VERY simple code-editing, and small modifications to your XX.organism.conf configuration file. These are described in detail below:

1 SYNOPSIS

In 0X.organism.conf, for example:

 [ORIGIN]
 link         = http://yoursite.com/cgi-bin/gbrowse_moby?source=$source&name=$name&class=$class&method=$method&ref=$ref&description=$description
 feature      = origin:Sequence
 glyph        = anchored_arrow
 fgcolor      = orange
 font2color   = red
 linewidth    = 2
 height       = 10
 description  = 1
 key          = Definition line
 link_target  = _MOBY

AND/OR

 [db_xref:DETAILS]
 URL = http://yoursite.com/cgi-bin/gbrowse_moby?namespace=$tag;id=$value

Note that all you are doing in each case is to associate a mouse click on a particular feature type with an invocation of the gbrowse_moby script, passing a few of the common Gbrowse variables in the GET string.

The gbrowse_moby script will take information passed from a click on a Gbrowse feature, or a click on a configured DETAILS GFF attribute type, and initiate a MOBY browsing session with information from that link. Most information is discarded. The only useful information to MOBY is a "namespace" and an "id" within that namespace.

Generally speaking, namespaces in Gbrowse will have to be mapped to a namespace in the MOBY namespace ontology (which is derived from the Gene Ontology Database Cross-Reference Abbreviations list). Currently, this requires editing of the gbrowse_moby code, where a Perl hash named %source2namespace maps the GFF source (column 2) to a MOBY namespace:

  $source2namespace{$source} = moby_namespace
2 REQUIRED LIBRARIES

This script requires libraries from the BioMOBY project. Currently these are only available from the CVS. Anonymous checkout of the BioMOBY project can be accomplished as follows:

  cvs -d :pserver:cvs@cvs.open-bio.org:/home/repository/moby login

When prompted for a password, type "cvs".

  cvs -d :pserver:cvs@cvs.open-bio.org:/home/repository/moby co moby-live
  cvs update -dP

You will then need to enter the moby-live/Perl folder and run "perl Makefile.PL; make; make install" to install the MOBY libraries into your system.

3 USAGE

gbrowse_moby understands the following variables, some of which (*) may be passed from Gbrowse through a mouse-click into the GET string:

 * $source    - converted into a MOBY namespace by parsing
              the 'source' GFF tag against the %source2namespace
              hash.
             (see more detailed explanation in the examples below)
 $namespace - used verbatim as a valid MOBY namespace
 * $name      - used verbatim as a MOBY id interpreted in the namespace
 * $id        - used verbatim as a MOBY id interpreted in the namespace
 * $class     - this is the GFF column 9 class; used for the page title
 $objectclass - this should be a MOBY Class ontology term
               (becomes Class 'Object' by default, and this
                is usually correct)
 $object      - contains the raw XML of a valid MOBY object 

Note that you MUST at least pass a namespace-type variable (source/namespace) and an id-type variable (name/id) in order to have a successful MOBY call.

4 EXAMPLES
Simple GFF

If your GFF were:

      A22344  Genbank  origin  1000  2000  87  +  .

You would set your configuration file as follows:

     [ORIGIN]
     link         = http://yoursite.com/cgi-bin/gbrowse_moby?source=$source&name=$name&class=$class
     feature      = origin:Genbank

and you would edit the gbrowse_moby script as follows:

      my %source2namespace = (
         #   GFF-source           MOBY-namespace
            'Genbank'       =>      'NCBI_Acc',
      );

this maps the GFF source tag "Genbank" to the MOBY namespace "NCBI_Acc"

GFF With non-MOBY Attributes

If your GFF were:

      A22344  Genbank origin  1000  2000 87 + . Locus CDC23

You would set your configuration file as follows:

     [ORIGIN]
     link         = http://yoursite.com/cgi-bin/gbrowse_moby?source=$source&name=$name&class=$class
     feature      = origin:Genbank

and you might also set a DETAILS call to handle the Locus Xref: (notice that we use the 'source' tag to force a translation of the foreign namespace into a MOBY namespace)

     [db_xref:DETAILS]
     URL = http://brie4.cshl.org:9320/cgi-bin/gbrowse_moby?source=$tag;id=$value

then to handle the mapping of Locus to YDB_Locus as well as the Genbank GFF source tag you would edit the source2namespace hash in gbrowse_moby to read:

      my %source2namespace = (
         #   GFF-source           MOBY-namespace
            'Genbank'       =>      'NCBI_Acc',
            'Locus'         =>      'YDB_Locus',
      );
GFF With MOBY Attributes

If your GFF were (NCBI_gi is a valid MOBY namespace):

      A22344  Genbank origin  1000  2000 87 + . NCBI_gi 118746

You would set your configuration file as follows:

     [ORIGIN]
     link         = http://yoursite.com/cgi-bin/gbrowse_moby?source=$source&name=$name&class=$class
     feature      = origin:Genbank

and you might also set a DETAILS call to handle the NCBI_gi Xref: (notice that we now use the 'namespace' tag to indicate that the tag is already a valid MOBY namespace)

     [db_xref:DETAILS]
     URL = http://brie4.cshl.org:9320/cgi-bin/gbrowse_moby?namespace=$tag;id=$value

Since there is no need to map the namespace portion, we now only need to handle the Genbank GFF source as before:

      my %source2namespace = (
         #   GFF-source           MOBY-namespace
            'Genbank'       =>      'NCBI_Acc',
      );
5 HINTS

-The full listing of valid MOBY namespaces is available at:

    http://mobycentral.cbr.nrc.ca/cgi-bin/types/Namespaces

-A useful mapping to make is to put the organism name into the Global_Keyword namespace. This will trigger discovery of MedLine searches for papers about that organism.

J. BioMOBY Services ^

A selection of services are distributed with the Gbrowse package that will allow you to serve your underlying data using the BioMOBY Services architecture.

To enable these, simply do the following:

1. Set-up and fill your database

as per the normal Gbrowse instructions

2. Edit the moby.conf file

in the /$CONFIG/gbrowse.conf/MobyServices folder. It should be set up as follows:

a. Reference

Your reference sequences will be based on some type of identifier - e.g. they will be from Genbank or from Embl or from Flybase, etc. Look-up the BioMOBY namespace corresponding to the type of identifier you are using for your Reference sequences and put that identifier here.

-The full listing of valid MOBY namespaces is available at:

    http://mobycentral.cbr.nrc.ca/cgi-bin/types/Namespaces
b. authURI

You are required to identify yourself when registering MOBY Services. Your authURI is a URI uniquely identifying you. This is generally your domain (e.g. flybase.org)

c. contactEmail

You are required to provide a contact email address to which people can contact you v.v. the services you are providing.

d. CGI_URL

This is simply the URL to the folder from which you are serving your gbrowse scripts. e.g. http://flybase.org/cgi-bin/browser/ DO NOT include the script name in this parameter! It is the folder only!!

e. [Namespace_Class_Mappings]

This section is just a list of tuples indicating the relationship between various entities in your database (e.g. Genes, Transcripts) and their equivalent BioMOBY namespaces. For example, if you are TAIR, and you have entities in your database called "Locus", you would add the line:

        Locus = TAIR_Locus

to this section of the config file. This will allow people who have TAIR_Locus identifiers in-hand to discover your service and request information about that locus from your database.

You may add as many Namespace->Class mappings as you wish; one per line.

3. REGISTERING SERVICES

To register your services with the MOBY Central web service registry simply run the "register_moby_services.pl" script, located in the Generic-Genome-Browser/bin folder. The script documentation can be retrieved with POD or simple documentation can be printed by simply running the script with no command-line parameters. Generally speaking you need only run:

perl register_moby_services.pl -register

As services are registered they will be added to a file: registeredMOBYServices.dat. This file is used to de-register your services if you wish to do so. To deregister, simply run:

perl register_moby_services.pl -clean

If your .dat file is not available, cleaning your services will be unsuccessful.

4. Service script

Your services are served by the script 'moby_server' in your cgi-bin folder. This is auto-configured by the register_services step above, so generally speaking you do not need to edit this script.

K. FILTERING SEARCH RESULTS ^

GBrowse provides a method to filter the contents of individual tracks based on information that can be obtained from feature attributes. For example, suppose you have performed a blast and added all hits as similarity features on an entry. In gbrowse, all those features can get a little crowdy. The administrator can decide to show only the top 5 of the blast hits. This can easily be accomplished by adding the filter option in the conf file. It might look like this:

  [BLAST]
  feature       = blast
  glyph         = segments
  filter = sub {
                 my $feat = shift;
                 (my $rank) = $feat->get_tag_values('rank'); # persistent Bio::SeqFeature::Generic features
                 #(my $rank) = $feat->attributes('rank'); # Bio::DB::GFF::Feature
                 $rank < 6;
               }

Another useful example is to show features coming from a plain genbank file. When loaded into BioSQL the source becomes 'EMBL/Genbank/SwissProt'. Using the Bio::DB::Das::BioSQL adaptor you have to pass the source to the feature option. It can be rather difficult to distinguish all the features when they all have the same source string. This problem can be solved using the filter option. In the following example the difference between the features is done based on the primary_tag

  [REGION]
  feature      = EMBL/GenBank/SwissProt
  filter       = sub {
                  my $feat = shift;
                  $feat->primary_tag =~ /region/i;
                 }
  key          = RefSeq Protein Domains
                                                                                
  [SIGPEPTIDE]
  feature      = EMBL/GenBank/SwissProt
  filter       = sub {
                  my $feat = shift;
                  $feat->primary_tag =~ /sig_peptide/i;
                 }
  key          = RefSeq Signal Peptide

L. INVOKING GBROWSE URLs (under construction) ^

This section describes the public CGI parameters recognized by GBrowse. By setting the parameters in the URL, you can get gbrowse to do various useful things:

The source argument

The last component of the gbrowse path is the symbolic name of the data source. For example:

   http://www.your.site/cgi-bin/gbrowse/volvox
   http://www.your.site/cgi-bin/gbrowse/yeast
   http://www.your.site/cgi-bin/gbrowse/my_testing_database

These will correspond to config files named volvox.pm, yeast.pm and my_testing_database.pm respectively.

As noted earlier, you can place numbers in front of the configuration file names in order to adjust the order in which they appear in the data source menu.

NOTE: For obscure reasons involving Internet Explorer compatibility, gbrowse will add an extra slash to the end of the URL, resulting in URLs that look like:

  http://www.your.site/cgi-bin/gbrowse/yeast/?q=NAB2

Don't worry about this. The URL works the same with and without the terminal slash.

q

The argument "q" will set the landmark or search string:

    http://www.your.site/cgi-bin/gbrowse/yeast?q=NAB2

This will have the same effect as typing "NAB2" into the gbrowse search box.

To go immediately to the multiple hits page (which shows hits on several overview panels), use multiple q arguments:

   http://www.your.site/cgi-bin/gbrowse/yeast?q=NAB2;q=NPY1

Alternatively, you can use a single q parameter and separate each landmark name with a dash:

   http://www.your.site/cgi-bin/gbrowse/yeast?q=NAB2-NPY1

The rules for specifying relative offsets and object classes are the same as in the main search field:

   http://www.your.site/cgi-bin/gbrowse/yeast?q=Gene:NAB2:1..5000
ref, start, stop, end

Together the "ref," "start" and "stop" arguments specify the reference sequence and the start and end coordinates of the region of interest. The "q" argument, if present, overrides these settings.

The "end" argument is a synonym for "stop".

label

The tracks to display. This parameter must contain the track names (i.e. the names in [brackets] in the config file) separated by "+" or "-" characters. For example:

   http://www.your.site/cgi-bin/gbrowse/yeast?labels=ORFs-tRNAs

To use the "+" character you may have to URL escape it:

   http://www.your.site/cgi-bin/gbrowse/yeast?labels=ORFs%2BtRNAs

All tracks not explicitly given by the label parameter will be closed (disabled).

enable

Tracks to enable. The tracks indicated by this parameter will be opened in addition to any tracks that were previously opened by the user. The format is the same as label:

   http://www.your.site/cgi-bin/gbrowse/yeast?enable=ORFs-tRNAs
disable

Tracks to close. The tracks indicated by this parameter will be disabled. Tracks not mentioned by this parameter will keep their previous state. The format is the same as label:

   http://www.your.site/cgi-bin/gbrowse/yeast?disable=ORFs-tRNAs

When modifying track state, the "label" parameter is processed first, followed by the "enable" parameter and the "disable" parameter.

flip

Whether to flip the display. If set to a true value (flip=1), then the coordinates will be reversed so that forward strand features become reverse strand features. If set to a false value (flip=0) or absent, then the forward strand is displayed as per usual.

width

Set the width of the overview, region and details images, in pixels.

region_size

Set the length of the region covered by the "region" panel, in base pairs.

add

A feature to superimpose on top of the current image as a new track. Multiple add arguments are allowed. The format is:

  reference+type+name+start..stop,start..stop,start..stop

example:

  add=chr3+EST+yk802.1+500000..500010,500100..500180
h_feat

The name of a feature to highlight in the format "<feature_name>@<color_name>". Example:

      h_feat=SKT5@blue

You may omit "@color", in which case the highlight will default to yellow. You can specify multiple h_feat arguments in order to highlight several features with distinct colors.

Passing an argument of h_feature=_clear_ will clear all feature highlighting.

h_region

The name of a region to highlight in the format "<seq_id>:start..end@color". Example:

      h_region=Chr3:200000..250000@wheat

You may omit "@color" in which case the highlight will default to lightgrey. You can specify multiple h_region arguments in order to highlight multiple sequence ranges with different colors.

Passing an argument of h_region=_clear_ will clear all region highlighting.

ks

The position of the key in the detail panel. Possible values are "between," "beneath," "left" and "right".

sk

The sort order of track names in the "Tracks" panel. Values are "sorted" (alphabetically sorted by name) and "unsorted" (sorted by the order of tracks as defined in the config file).

track_options

If true, open up the track configuration page.

help

Open up the specified help page. Possible values are:

     "general"    open the general help page
     "citations"  open up the track description & citation page
     "link_image" open the page that describes how to
                  generate an embedded image of the current view
      "svg_image" the page that describes how to generate SVGs
add

Upload a feature and add it in its own track. The format is "reference+type+name+start..end", where reference is the landmark for the coordinates (e.g. a named gene or chromosome), type is the type of the feature, name is the name of the feature, and start..end are the start and end coordinates. For a feature that has multiple segments, you may use multiple start..end ranges, separated by commas. Example:

  add=chr3+miRNA+mir144+2309229..2309300,2309501..2309589

Pass multiple "add" parameters to upload several features.

"add" can be abbreviated to "a" for terseness.

style

Specify the style for features uploaded using "add". It is a flattened version of the style configuration sections described in this document. Lines are separated by "+" symbols rather than newlines. The first word in the argument is the feature type to configure, for example "miRNA." Subsequent option=value pairs control the glyph and glyph options.

For example, if you have added a "miRNA" annotation, then you can tell the renderer to use a red arrow for this glyph in this way:

   style=miRNA+glyph=arrow+fgcolor=red

"style" can be abbreviated to "s" for terseness.

id

The id is a unique session ID that will store persistent configuration information. You do not typically need to use the id parameter except in the circumstance in which you wish to upload an annotation file programatically, in which case you should choose some large hard-to-guess number.

Upload, upload_annotations, id

These three arguments must be present in order to upload a file of external annotations to the server. "Upload" must be a true value (such as "1"), and "upload_annotations" will contain the content of the uploaded file. Note that you must POST the data using MIME type "multipart/form-data" and that the "U" in upload is capitalized.

The "id" argument is used to associated the upload with a session. Pick some long, hard to guess number. This will be associated stably with the uploaded file(s). To see the upload information, provide the same number in the "id" argument every time you access gbrowse.

eurl

Specify the URL of a remote annotation source to load into the database. You should also supply an "id" argument as well, as described earlier, in order to be able to view the annotations.

plugin, plugin_do

These arguments run plugins. The "plugin" argument gives the name of the plugin to activate. The name is the last component of the plugin package name, e.g. FastaDumper. The "plugin_do" argument selects what to do with the plugin. Possible values are "Configure", "Find" and "Go". "Configure" launches the plugin's configure page, "Go" runs dumper plugins' dump operation, and "Find" activates finder plugins' find function. For find operations, you should in most cases pass the find string in the "q" argument, but this depends on the particular plugin.

Each plugin may have its own set of URL arguments. A plugin's arguments are preceded by the plugin's name. For example, the FastaDumper plugin has a parameter named "format" which controls the output format. So to invoke this plugin and make the output plain text, one would provide the arguments:

 http://www.your.site/cgi-bin/gbrowse/yeast?q=NUT21;plugin=FastaDumper;
             plugin_do=Go;FastaDumper.format=text

Plugins tend not to be well documented, so you may have to read through the source code to figure out their arguments.

M. FURTHER INFORMATION ^

For further information, bug reports, etc, please consult the mailing lists at www.gmod.org. The main mailing list for gbrowse support is gmod-gbrowse@lists.sourceforge.net.

Have fun!

Lincoln Stein & the GMOD development team lstein@cshl.edu

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