GMOD

GMOD Online Training 2014/WebApollo Tutorial

Contents

Using WebApollo

We’ve made some configuration modifications after the original AMI was created, so run the following commands in your VM:

$ cd
$ wget icebox.lbl.gov/webapollo/gmod/2014/summer_school/new_config.tgz
$ cd /var/lib/tomcat7/webapps/WebApolloDemo/config
$ tar -xvzf ~/new_config.tgz
$ sudo service tomcat7 restart

WebApollo is a web-based application, so the only requirement to use it is a web browser. It has been tested with Chrome, Firefox, and Safari. It has not been tested with Internet Explorer.

A WebApollo demo with the Pythium data has been set up on the virtual machine. We’ll use the demo to view WebApollo’s functionality.

Point your browser to http://ec2-##-##-##-##.compute-1.amazonaws.com/WebApolloDemo.
(e.g., http://ec2-184-73-92-243.compute-1.amazonaws.com/WebApolloDemo).

WebApollo login page

The user name and password are both demo.

WebApollo reference sequence selection

We only have one contig to work with. Click on the ` scf1117875582023` link.

Installing WebApollo

Server operating system

Any Unix like system (e.g., Unix, Linux, Mac OS X)

Prerequisites

Note: All prerequisites have already been installed on the course’s machine

Installation

The installation steps will be done in the command line. SSH into your AWS machine. Uncompress the WebApollo-2014-04-03.tgz tarball.

$ cd
$ tar -xvzf sources/WebApollo-2014-04-03.tgz

User database

WebApollo uses a database to determine who can access and edit annotations for a given sequence.

First we’ll need to create a database. You can call it whatever you want (remember the name as you’ll need to point the configuration to it). For the purposes of this guide, we’ll call it web_apollo_users. You might want to create a separate account to manage the database. We’ll have the user web_apollo_db with password web_apollo_db. The user will not be a superuser nor will it be able to create new roles. But it will be able to create databases.

$ createuser -P web_apollo_db
Enter password for new role: 
Enter it again: 
Shall the new role be a superuser? (y/n) n
Shall the new role be allowed to create databases? (y/n) y
Shall the new role be allowed to create more new roles? (y/n) n

Next we’ll create the user database.

$ createdb -U web_apollo_db web_apollo_users

Now that the database is created, we need to load the schema to it.

$ cd ~/WebApollo-2014-04-03/tools/user
$ psql -U web_apollo_db web_apollo_users < user_database_postgresql.sql

Now the user database has been setup.

Let’s populate the database.

First we’ll create an user with access to WebApollo. We’ll use the add_user.pl script in ~/WebApollo-2014-04-03/tools/user. Let’s create an user named web_apollo_admin with the password web_apollo_admin.

$ ./add_user.pl -D web_apollo_users -U web_apollo_db -P web_apollo_db \
-u web_apollo_admin -p web_apollo_admin

Next we’ll add the annotation tracks ids for the genomic sequences for our organism. We’ll use the add_tracks.pl script in the same directory. We need to generate a file of genomic sequence ids for the script. For convenience, there’s a script called extract_seqids_from_fasta.pl in the same directory which will go through a FASTA file and extract all the ids from the deflines. Let’s first create the list of genomic sequence ids. We’ll store it in ~/scratch/seqids.txt. We’ll want to add the prefix “Annotations-” to each identifier.

$ mkdir ~/scratch
$ ./extract_seqids_from_fasta.pl -p Annotations- -i ~/sample_data/pyu_data/scf1117875582023.fa \
-o ~/scratch/seqids.txt

Now we’ll add those ids to the user database.

$ ./add_tracks.pl -D web_apollo_users -U web_apollo_db -P web_apollo_db \
-t ~/scratch/seqids.txt

Now that we have an user created and the annotation track ids loaded, we’ll need to give the user permissions to access the sequence. We’ll have the all permissions (read, write, publish, user manager). We’ll use the set_track_permissions.pl script in the same directory. We’ll need to provide the script a list of genomic sequence ids, like in the previous step.

$ ./set_track_permissions.pl -D web_apollo_users -U web_apollo_db \
-P web_apollo_db -u web_apollo_admin -t ~/scratch/seqids.txt -a

We’re all done setting up the user database.

Note that we’re only using a subset of the options for all the scripts mentioned above. You can get more detailed information on any given script (and other available options) using the “-h” or “–help” flag when running the script.

Deploying the servlet

Tomcat error handling

Note that WebApollo server sends error to the client through JSON messages. Your servlet container must be configured to allow raw JSON to be sent as when errors occur. In the case of Tomcat, you’ll need to configure it to use the custom valve that is provided with the WebApollo package. This step only needs to be done once per Tomcat server.

This step has already been done.

$ cp ~/WebApollo-2014-04-03/tomcat/custom-valves.jar /usr/share/tomcat7/lib

You’ll then need to add errorReportValveClass="org.bbop.apollo.web.ErrorReportValve" as an attribute to the existing <Host> element in /etc/tomcat7/server.xml

<Host name="localhost" appBase="webapps" 
      unpackWARs="true" autoDeploy="true" 
      errorReportValveClass="org.bbop.apollo.web.ErrorReportValve">
</Host>

Deploying the WAR

We need to deploy the WAR file in the war directory from the unpacked tarball.

$ cd /var/lib/tomcat7/webapps

Next we need to create the directory that will contain the application.

$ sudo mkdir WebApollo

Let’s make ubuntu the owner of the newly created directory, so that we don’t have to constantly run commands as root.

$ sudo chown ubuntu WebApollo

Now we’ll go into the newly created directory and unpack the WAR file into it.

$ cd WebApollo
$ jar -xvf ~/WebApollo-2014-04-03/war/WebApollo.war

That’s it! We’re done installing WebApollo. Now we need to move on to configuring the application.

Configuration

Most configuration files will reside in /var/lib/tomcat7/webapps/WebApollo/config. We’ll need to configure a number of things before we can get WebApollo up and running.

Supported annotation types

Many configurations will require you to define which annotation types the configuration will apply to. WebApollo supports the following “higher level” types (from the Sequence Ontology):

Main configuration

The main configuration is stored in /var/lib/tomcat7/webapps/WebApollo/config/config.xml. Let’s take a look at the file.

<?xml version="1.0" encoding="UTF-8"?>
<server_configuration>
 
    <!-- mapping configuration for GBOL data structures -->
    <gbol_mapping>/config/mapping.xml</gbol_mapping>
 
    <!-- directory where JE database will be created -->
    <datastore_directory>ENTER_DATASTORE_DIRECTORY_HERE</datastore_directory>
 
    <!-- minimum size for introns created -->
    <default_minimum_intron_size>1</default_minimum_intron_size>
 
    <!-- size of history for each feature - setting to 0 means unlimited history -->
    <history_size>0</history_size>
 
        <!-- overlapping strategy for adding transcripts to genes -->
        <overlapper_class>org.bbop.apollo.web.overlap.OrfOverlapper</overlapper_class>
 
        <!-- javascript file for comparing track names (refseqs) (used for sorting in selection table) -->
        <track_name_comparator>/config/track_name_comparator.js</track_name_comparator>
 
        <!-- whether to use an existing CDS when creating new transcripts -->
        <use_cds_for_new_transcripts>true</use_cds_for_new_transcripts>
 
    <!-- set to false to use hybrid disk/memory store which provides a little slower performance
 but uses a lot less memory - great for annotation rich genomes -->
    <use_pure_memory_store>true</use_pure_memory_store>
 
    <!-- user authentication/permission configuration -->
    <user>
 
        <!-- database configuration -->
        <database>
 
            <!-- driver for user database -->
            <driver>org.postgresql.Driver</driver>
 
            <!-- JDBC URL for user database -->
            <url>ENTER_USER_DATABASE_JDBC_URL</url>
 
            <!-- username for user database -->
            <username>ENTER_USER_DATABASE_USERNAME</username>
 
            <!-- password for user database -->
            <password>ENTER_USER_DATABASE_PASSWORD</password>
 
        </database>
 
        <!-- class for generating user authentication page
         (login page) -->
        <authentication_class>org.bbop.apollo.web.user.localdb.LocalDbUserAuthentication</authentication_class>
 
    </user>
 
    <tracks>
 
        <!-- path to JBrowse refSeqs.json file -->
        <refseqs>ENTER_PATH_TO_REFSEQS_JSON_FILE</refseqs>
 
        <!-- annotation track name the current convention is to append
         the genomic region id to the the name of the annotation track
         e.g., if the annotation track is called "Annotations" and the
         genomic region is chr2L, the track name will be
         "Annotations-chr2L".-->
        <annotation_track_name>Annotations</annotation_track_name>
 
        <!-- organism being annotated -->
        <organism>ENTER_ORGANISM</organism>
 
        <!-- CV term for the genomic sequences - should be in the form
         of "CV:term".  This applies to all sequences -->
        <sequence_type>ENTER_CVTERM_FOR_SEQUENCE</sequence_type>
 
        <!-- path to file containing translation table.
         optional - defaults to NCBI translation table 1 if absent -->
        <translation_table>/config/translation_tables/ncbi_1_translation_table.txt</translation_table>
 
        <!-- splice acceptor and donor sites. Multiple entries may be
         added to allow multiple accepted sites.
         optional - defaults to GT for donor and AG for acceptor
         if absent -->
        <splice_sites>
            <donor_site>GT</donor_site>
            <acceptor_site>AG</acceptor_site>
        </splice_sites>
 
    </tracks>
 
    <!-- path to file containing canned comments XML -->
    <canned_comments>/config/canned_comments.xml</canned_comments>
 
    <!-- configuration for what to display in the annotation info editor.
     Sections can be commented out to not be displayed or uncommented
     to make them active -->
    <annotation_info_editor>
 
        <!-- grouping for the configuration.  The "feature_types" attribute takes a list of
     SO terms (comma separated) to apply this configuration to
     (e.g., feature_types="sequence:transcript,sequence:mRNA" will make it so the group
     configuration will only apply to features of type "sequence:transcript" or "sequence:mRNA").
     A value of "default" will make this the default configuration for any types not explicitly
     defined in other groups.  You can have any many groups as you'd like -->
        <annotation_info_editor_group feature_types="default">
 
            <!-- display status section.  The text for each <status_flag>
         element will be displayed as a radio button in the status
         section, in the same order -->
            <!--
         <status>
             <status_flag>Approved</status_flag>
             <status_flag>Needs review</status_flag>
         </status>
         -->
 
            <!-- display generic attributes section -->
            <attributes />
 
            <!-- display dbxrefs section -->
            <dbxrefs />
 
            <!-- display PubMed IDs section -->
            <pubmed_ids />
 
            <!-- display GO IDs section -->
            <go_ids />
 
            <!-- display comments section -->
            <comments />
 
        </annotation_info_editor_group>
 
    </annotation_info_editor>
 
    <!-- tools to be used for sequence searching.  This is optional.
     If this is not setup, WebApollo will not have sequence search support -->
    <sequence_search_tools>
 
        <!-- one <sequence_search_tool> element per tool -->
        <sequence_search_tool>
 
            <!-- display name for the search tool -->
            <key>BLAT nucleotide</key>
 
            <!-- class for handling search -->
            <class>org.bbop.apollo.tools.seq.search.blat.BlatCommandLineNucleotideToNucleotide</class>
 
            <!-- configuration for search tool -->
            <config>/config/blat_config.xml</config>
 
        </sequence_search_tool>
 
        <sequence_search_tool>
 
            <!-- display name for the search tool -->
            <key>BLAT protein</key>
 
            <!-- class for handling search -->
            <class>org.bbop.apollo.tools.seq.search.blat.BlatCommandLineProteinToNucleotide</class>
 
            <!-- configuration for search tool -->
            <config>/config/blat_config.xml</config>
 
        </sequence_search_tool>
 
    </sequence_search_tools>
 
    <!-- data adapters for writing annotation data to different formats.
     These will be used to dynamically generate data adapters within
     WebApollo.  This is optional.  -->
    <data_adapters>
 
        <!-- one <data_adapter> element per data adapter -->
        <data_adapter>
 
            <!-- display name for data adapter -->
            <key>GFF3</key>
 
            <!-- class for data adapter plugin -->
            <class>org.bbop.apollo.web.dataadapter.gff3.Gff3DataAdapter</class>
 
            <!-- required permission for using data adapter
         available options are: read, write, publish -->
            <permission>read</permission>
 
            <!-- configuration file for data adapter -->
            <config>/config/gff3_config.xml</config>
 
            <!-- options to be passed to data adapter -->
            <options>output=file&amp;format=gzip</options>
 
        </data_adapter>
 
        <data_adapter>
 
            <!-- display name for data adapter -->
            <key>Chado</key>
 
            <!-- class for data adapter plugin -->
            <class>org.bbop.apollo.web.dataadapter.chado.ChadoDataAdapter</class>
 
            <!-- required permission for using data adapter
         available options are: read, write, publish -->
            <permission>publish</permission>
 
            <!-- configuration file for data adapter -->
            <config>/config/chado_config.xml</config>
 
            <!-- options to be passed to data adapter -->
            <options>display_features=false</options>
 
        </data_adapter>
 
        <!-- group the <data_adapter> children elements together -->
        <data_adapter_group>
 
            <!-- display name for adapter group -->
            <key>FASTA</key>
 
            <!-- required permission for using data adapter group
         available options are: read, write, publish -->
            <permission>read</permission>
 
            <!-- one child <data_adapter> for each data adapter in the group -->
            <data_adapter>
 
                <!-- display name for data adapter -->
                <key>peptide</key>
 
                <!-- class for data adapter plugin -->
                <class>org.bbop.apollo.web.dataadapter.fasta.FastaDataAdapter</class>
 
                <!-- required permission for using data adapter
             available options are: read, write, publish -->
                <permission>read</permission>
 
                <!-- configuration file for data adapter -->
                <config>/config/fasta_config.xml</config>
 
                <!-- options to be passed to data adapter -->
                <options>output=file&amp;format=gzip&amp;seqType=peptide</options>
 
            </data_adapter>
 
            <data_adapter>
 
                <!-- display name for data adapter -->
                <key>cDNA</key>
 
                <!-- class for data adapter plugin -->
                <class>org.bbop.apollo.web.dataadapter.fasta.FastaDataAdapter</class>
 
                <!-- required permission for using data adapter
             available options are: read, write, publish -->
                <permission>read</permission>
 
                <!-- configuration file for data adapter -->
                <config>/config/fasta_config.xml</config>
 
                <!-- options to be passed to data adapter -->
                <options>output=file&amp;format=gzip&amp;seqType=cdna</options>
 
            </data_adapter>
 
            <data_adapter>
 
                <!-- display name for data adapter -->
                <key>CDS</key>
 
                <!-- class for data adapter plugin -->
                <class>org.bbop.apollo.web.dataadapter.fasta.FastaDataAdapter</class>
 
                <!-- required permission for using data adapter
             available options are: read, write, publish -->
                <permission>read</permission>
 
                <!-- configuration file for data adapter -->
                <config>/config/fasta_config.xml</config>
 
                <!-- options to be passed to data adapter -->
                <options>output=file&amp;format=gzip&amp;seqType=cds</options>
 
            </data_adapter>
 
        </data_adapter_group>
 
    </data_adapters>
 
</server_configuration>

Let’s look through each element in more detail with values filled in.

<!-- mapping configuration for GBOL data structures -->
<gbol_mapping>/config/mapping.xml</gbol_mapping>

File that contains type mappings used by the underlying data model. It’s best not to change the default option.

<!-- directory where JE database will be created -->
<datastore_directory>/data/webapollo/annotations</datastore_directory>

Directory where user generated annotations will be stored. The data is stored using Berkeley DB. We’ll use /data/webapollo/annotations.

<!-- minimum size for introns created -->
<default_minimum_intron_size>1</default_minimum_intron_size>

Minimum length of intron to be created when using the “Make intron” operation. The operation will try to make the shortest intron that’s at least as long as this parameter. So if you set it to a value of “40”, then all calculated introns will be at least of length 40.

<!-- size of history for each feature - setting to 0 means unlimited history -->
<history_size>0</history_size>

The size of your history stack, meaning how many “Undo/Redo” steps you can do. The larger the number, the larger the storage space needed. Setting it to “0” makes it to that there’s no limit.

<!-- overlapping strategy for adding transcripts to genes -->
<overlapper_class>org.bbop.apollo.web.overlap.OrfOverlapper</overlapper_class>

Defines the strategy to be used for deciding whether overlapping transcripts should be considered splice variants to the same gene. This points to a Java class implementing the org.bbop.apollo.overlap.Overlapper interface. This allows you to create your own custom overlapping strategy should the need arise. Currently available options are:

<!-- javascript file for comparing track names (refseqs) (used for sorting in selection table) -->
<track_name_comparator>/config/track_name_comparator.js</track_name_comparator>

Defines how to compare genomic sequence names for sorting purposes in the genomic region selection list. Points to a javascript file. You can implement your logic to allow whatever sorting you’d like for your own organism. This doesn’t make much of a difference in our case since we’re only dealing with one genomic region. The default behavior is to sort names lexicographically.

<!-- whether to use an existing CDS when creating new transcripts -->
<use_cds_for_new_transcripts>true</use_cds_for_new_transcripts>

Tells WebApollo whether to use an existing CDS when creating a new transcript (otherwise it computes the longest ORF). This can be useful when gene predictors suggest a CDS that’s not the longest ORF and you want to use that instead. This is only applicable when using features that have a CDS associated with them.

<!-- set to false to use hybrid disk/memory store which provides a little slower performance
but uses a lot less memory - great for annotation rich genomes -->
<use_pure_memory_store>true</use_pure_memory_store>

Defines whether the internal data store is purely a memory one or a hybrid memory/disk store. The memory store provides faster performance at the cost of more memory. The hybrid store provides a little slower performance but uses a lot less memory, so it’s a good option for annotation rich genomes. Set to true to use the memory store and false to use the hybrid one.

Let’s take look at the user element, which handles configuration for user authentication and permission handling.

<!-- user authentication/permission configuration -->
<user>
 
    <!-- database configuration -->
    <database>
 
        <!-- driver for user database -->
        <driver>org.postgresql.Driver</driver>
 
        <!-- JDBC URL for user database -->
        <url>ENTER_USER_DATABASE_JDBC_URL</url>
 
        <!-- username for user database -->
        <username>ENTER_USER_DATABASE_USERNAME</username>
 
        <!-- password for user database -->
        <password>ENTER_USER_DATABASE_PASSWORD</password>
 
    </database>
 
    <!-- class for generating user authentication page (login page) -->
    <authentication_class>org.bbop.apollo.web.user.localdb.LocalDbUserAuthentication</authentication_class>
 
</user>

Let’s first look at the database element that defines the database that will handle user permissions (which we created previously).

<!-- driver for user database -->
<driver>org.postgresql.Driver</driver>

This should point the JDBC driver for communicating with the database. We’re using a PostgreSQL driver since that’s the database we’re using for user permission management.

<!-- JDBC URL for user database -->
<url>jdbc:postgresql://localhost/web_apollo_users</url>

JDBC URL to the user permission database. We’ll use jdbc:postgresql://localhost/web_apollo_users since the database is running in the same server as the annotation editing engine and we named the database web_apollo_users.

<!-- username for user database -->
<username>web_apollo_db</username>

User name that has read/write access to the user database. The user with access to the user database has the user name web_apollo_db.

<!-- password for user database -->
<password>web_apollo_db</password>

Password to access user database. The user with access to the user database has the password </tt>web_apollo_db</tt>.

Now let’s look at the other elements in the user element.

<!-- class for generating user authentication page (login page) -->
<authentication_class>org.bbop.apollo.web.user.localdb.LocalDbUserAuthentication</authentication_class>

Defines how user authentication is handled. This points to a class implementing the org.bbop.apollo.web.user.UserAuthentication interface. This allows you to implement any type of authentication you’d like (e.g., LDAP). Currently available options are:

Now let’s look at the configuration for accessing the annotation tracks for the genomic sequences.

<tracks>
 
    <!-- path to JBrowse refSeqs.json file -->
    <refseqs>ENTER_PATH_TO_REFSEQS_JSON_FILE</refseqs>
 
    <!-- annotation track name the current convention is to append
     the genomic region id to the the name of the annotation track
     e.g., if the annotation track is called "Annotations" and the
     genomic region is chr2L, the track name will be
     "Annotations-chr2L".-->
    <annotation_track_name>Annotations</annotation_track_name>
 
    <!-- organism being annotated -->
    <organism>ENTER_ORGANISM</organism>
 
    <!-- CV term for the genomic sequences - should be in the form
     of "CV:term".  This applies to all sequences -->
    <sequence_type>ENTER_CVTERM_FOR_SEQUENCE</sequence_type>
 
    <!-- path to file containing translation table.
     optional - defaults to NCBI translation table 1 if absent -->
    <translation_table>/config/translation_tables/ncbi_1_translation_table.txt</translation_table>
 
    <!-- splice acceptor and donor sites. Multiple entries may be
     added to allow multiple accepted sites.
     optional - defaults to GT for donor and AG for acceptor
     if absent -->
    <splice_sites>
        <donor_site>GT</donor_site>
        <acceptor_site>AG</acceptor_site>
    </splice_sites>
 
</tracks>

Let’s look at each element individually.

<!-- path to JBrowse refSeqs.json file -->
<refseqs>/var/lib/tomcat7/webapps/WebApollo/jbrowse/data/seq/refSeqs.json</refseqs>

Location where the refSeqs.json file resides, which is created from the data generation pipeline (see the data generation section). By default, the JBrowse data needs to reside in /var/lib/tomcat7/webapps/WebApollo/jbrowse/data. If you want the data to reside elsewhere, you’ll need to do configure your servlet container to handle the appropriate alias to jbrowse/data or symlink the data directory to somewhere else. WebApollo is pre-configured to allow symlinks.

<annotation_track_name>Annotations</annotation_track_name>

Name of the annotation track. Leave it as the default value of Annotations.

<!-- organism being annotated -->
<organism>Pythium ultimum</organism>

Scientific name of the organism being annotated (genus and species). We’re annotating Pythium ultimum.

<!-- CV term for the genomic sequences - should be in the form
 of "CV:term".  This applies to all sequences -->
<sequence_type>sequence:contig</sequence_type>

The type for the genomic sequences. Should be in the form of CV:term. Our genomic sequences are of the type sequence:contig.

<!-- path to file containing translation table.
 optional - defaults to NCBI translation table 1 if absent -->
<translation_table>/config/translation_tables/ncbi_1_translation_table.txt</translation_table>

File that contains the codon translation table. This is optional and defaults to NCBI translation table 1 if absent. See the translation tables section for details on which tables are available and how to customize your own table.

<!-- splice acceptor and donor sites. Multiple entries may be
 added to allow multiple accepted sites.
 optional - defaults to GT for donor and AG for acceptor
 if absent -->
<splice_sites>
    <donor_site>GT</donor_site>
    <acceptor_site>AG</acceptor_site>
</splice_sites>

Defines what the accepted donor and acceptor splice sites are. This will determine whether the client displays a warning on splice sites (if the splice site sequence doesn’t match what’s defined here, then it flags the splice site). You can add multiple <donor_site> and <acceptor_site> elements if your organism should support multiple values. This is optional and defaults to GT for donor and AG for acceptor sites.

<!-- path to file containing canned comments XML -->
<canned_comments>/config/canned_comments.xml</canned_comments>

File that contains canned comments (predefined comments that will be available from a pull-down menu when creating comments). It’s best not to change the default option. See the canned comments section for details on configuring canned comments.

<!-- configuration for what to display in the annotation info editor.
Sections can be commented out to not be displayed or uncommented
to make them active -->
<annotation_info_editor>
 
    <!-- grouping for the configuration.  The "feature_types" attribute takes a list of
 SO terms (comma separated) to apply this configuration to
 (e.g., feature_types="sequence:transcript,sequence:mRNA" will make it so the group
 configuration will only apply to features of type "sequence:transcript" or "sequence:mRNA").
 A value of "default" will make this the default configuration for any types not explicitly
 defined in other groups.  You can have any many groups as you'd like -->
    <annotation_info_editor_group feature_types="default">
 
        <!-- display status section.  The text for each <status_flag>
     element will be displayed as a radio button in the status
     section, in the same order -->
        <!--
     <status>
         <status_flag>Approved</status_flag>
         <status_flag>Needs review</status_flag>
     </status>
     -->
 
        <!-- display generic attributes section -->
        <attributes />
 
        <!-- display dbxrefs section -->
        <dbxrefs />
 
        <!-- display PubMed IDs section -->
        <pubmed_ids />
 
        <!-- display GO IDs section -->
        <go_ids />
 
        <!-- display comments section -->
        <comments />
 
    </annotation_info_editor_group>
 
</annotation_info_editor>

Here’s the configuration on what to display in the annotation info editor. It will always display Name, Symbol, and Description but the rest is optional. This allows you to make the editor more compact if you’re not interested in editing certain metadata. Let’s look at the options in more detail.

<!-- grouping for the configuration.  The "feature_types" attribute takes a list of
SO terms (comma separated) to apply this configuration to
(e.g., feature_types="sequence:transcript,sequence:mRNA" will make it so the group
configuration will only apply to features of type "sequence:transcript" or "sequence:mRNA").
A value of "default" will make this the default configuration for any types not explicitly
defined in other groups.  You can have any many groups as you'd like -->
<annotation_info_editor_group feature_types="default">
    ...
</annotation_info_editor_group>

Each configuration is grouped by annotation type. This allows you to have different options on what’s displayed for specified types. The feature_types attribute defines which types this group will apply to. feature_types takes a list of SO terms (comma separated), such as "sequence:transcript,sequence:mRNA", which will apply this configuration to annotations of type sequence:transcript and sequence:mRNA. Alternatively, you can set the value to "default" which will become the default configuration for any types not explicitly defined in other groups. You can have any many groups as you’d like. All supported annotation types can be used.

Next, let’s look at each item to configure in each group.

<!-- display status section.  The text for each <status_flag>
 element will be displayed as a radio button in the status
 section, in the same order -->
<status>
    <status_flag>Approved</status_flag>
    <status_flag>Needs review</status_flag>
</status>

Allows selecting the status for a particular annotation. The value for <status_flag> is arbitrary (you can enter any text) and you can add as many as you’d like, but you need to at least have one (they’ll show up as selectable buttons in the editor).

<!-- display generic attributes section -->
<attributes />

Allows editing of generic attributes (tag/value pairs). Think non-reserved GFF3 tags for column 9.

<!-- display dbxrefs section -->
<dbxrefs />

Allows editing of database cross references.

<!-- display PubMed IDs section -->
<pubmed_ids />

Allows editing of PubMed IDs (for associating an annotation with a publication).

<!-- display GO IDs section -->
<go_ids />

Allows editing of Gene Ontology terms (for associating an annotation to a particular function).

<!-- display comments section -->
<comments />

Allows editing of comments for annotations.

<!-- tools to be used for sequence searching.  This is optional.
 If this is not setup, WebApollo will not have sequence search support -->
<sequence_search_tools>
 
    <!-- one <sequence_search_tool> element per tool -->
    <sequence_search_tool>
 
        <!-- display name for the search tool -->
        <key>BLAT nucleotide</key>
 
        <!-- class for handling search -->
        <class>org.bbop.apollo.tools.seq.search.blat.BlatCommandLineNucleotideToNucleotide</class>
 
        <!-- configuration for search tool -->
        <config>/config/blat_config.xml</config>
 
    </sequence_search_tool>
 
    <sequence_search_tool>
 
        <!-- display name for the search tool -->
        <key>BLAT protein</key>
 
        <!-- class for handling search -->
        <class>org.bbop.apollo.tools.seq.search.blat.BlatCommandLineProteinToNucleotide</class>
 
        <!-- configuration for search tool -->
        <config>/config/blat_config.xml</config>
 
    </sequence_search_tool>
 
</sequence_search_tools>

Here’s the configuration for sequence search tools (allows searching your genomic sequences). WebApollo does not implement any search algorithms, but instead relies on different tools and resources to handle searching (this provides much more flexible search options). This is optional. If it’s not configured, WebApollo will not have sequence search support. You’ll need one sequence_search_tool element per search tool. Let’s look at the element in more detail.

<!-- display name for the search tool -->
<key>BLAT nucleotide</key>

This is a string that will be used for the display name for the search tool, in the pull down menu that provides search selection for the user.

<!-- class for handling search -->
<class>org.bbop.apollo.tools.seq.search.blat.BlatCommandLineNucleotideToNucleotide</class>

Should point to the class that will handle the search request. Searching is handled by classes that implement the org.bbop.apollo.tools.seq.search.SequenceSearchTool interface. This allows you to add support for your own favorite search tools (or resources). We currently only have support for command line Blat, in the following flavors:

<!-- configuration for search tool -->
<config>/config/blat_config.xml</config>

File that contains the configuration for the searching plugin chosen. If you’re using Blat, you should not change this. If you’re using your own plugin, you’ll want to point this to the right configuration file (which will be dependent on your plugin). See the Blat section for details on configuring WebApollo to use Blat.

<!-- data adapters for writing annotation data to different formats.
These will be used to dynamically generate data adapters within
WebApollo.  It contains either <data_adapter> or <data_adapter_group> elements.
<data_adapter_group> will allow grouping adapters together and will provide a
submenu for those adapters in WebApollo. This is optional.  -->
<data_adapters>
 
    <!-- one <data_adapter> element per data adapter -->
    <data_adapter>
 
        <!-- display name for data adapter -->
        <key>GFF3</key>
 
        <!-- class for data adapter plugin -->
        <class>org.bbop.apollo.web.dataadapter.gff3.Gff3DataAdapter</class>
 
        <!-- required permission for using data adapter
     available options are: read, write, publish -->
        <permission>read</permission>
 
        <!-- configuration file for data adapter -->
        <config>/config/gff3_config.xml</config>
 
        <!-- options to be passed to data adapter -->
        <options>output=file&amp;format=gzip</options>
 
    </data_adapter>
 
    <data_adapter>
 
        <!-- display name for data adapter -->
        <key>Chado</key>
 
        <!-- class for data adapter plugin -->
        <class>org.bbop.apollo.web.dataadapter.chado.ChadoDataAdapter</class>
 
        <!-- required permission for using data adapter
     available options are: read, write, publish -->
        <permission>publish</permission>
 
        <!-- configuration file for data adapter -->
        <config>/config/chado_config.xml</config>
 
        <!-- options to be passed to data adapter -->
        <options>display_features=false</options>
 
    </data_adapter>
 
    <!-- group the <data_adapter> children elements together -->
    <data_adapter_group>
 
        <!-- display name for adapter group -->
        <key>FASTA</key>
 
        <!-- required permission for using data adapter group
     available options are: read, write, publish -->
        <permission>read</permission>
 
        <!-- one child <data_adapter> for each data adapter in the group -->
        <data_adapter>
 
            <!-- display name for data adapter -->
            <key>peptide</key>
 
            <!-- class for data adapter plugin -->
            <class>org.bbop.apollo.web.dataadapter.fasta.FastaDataAdapter</class>
 
            <!-- required permission for using data adapter
         available options are: read, write, publish -->
            <permission>read</permission>
 
            <!-- configuration file for data adapter -->
            <config>/config/fasta_config.xml</config>
 
            <!-- options to be passed to data adapter -->
            <options>output=file&amp;format=gzip&amp;seqType=peptide</options>
 
        </data_adapter>
 
        <data_adapter>
 
            <!-- display name for data adapter -->
            <key>cDNA</key>
 
            <!-- class for data adapter plugin -->
            <class>org.bbop.apollo.web.dataadapter.fasta.FastaDataAdapter</class>
 
            <!-- required permission for using data adapter
         available options are: read, write, publish -->
            <permission>read</permission>
 
            <!-- configuration file for data adapter -->
            <config>/config/fasta_config.xml</config>
 
            <!-- options to be passed to data adapter -->
            <options>output=file&amp;format=gzip&amp;seqType=cdna</options>
 
        </data_adapter>
 
        <data_adapter>
 
            <!-- display name for data adapter -->
            <key>CDS</key>
 
            <!-- class for data adapter plugin -->
            <class>org.bbop.apollo.web.dataadapter.fasta.FastaDataAdapter</class>
 
            <!-- required permission for using data adapter
         available options are: read, write, publish -->
            <permission>read</permission>
 
            <!-- configuration file for data adapter -->
            <config>/config/fasta_config.xml</config>
 
            <!-- options to be passed to data adapter -->
            <options>output=file&amp;format=gzip&amp;seqType=cds</options>
 
        </data_adapter>
 
    </data_adapter_group>
 
</data_adapters>

Here’s the configuration for data adapters (allows writing annotations to different formats). This is optional. If it’s not configured, WebApollo will not have data writing support. You’ll need one <data_adapter> element per data adapter. You can group data adapters by placing each <data_adapter> inside a <data_adapter_group> element. Let’s look at the <data_adapter> element in more detail.

<!-- display name for data adapter -->
<key>GFF3</key>

This is a string that will be used for the data adapter name, in the dynamically generated data adapters list for the user.

<!-- class for data adapter plugin -->
<class>org.bbop.apollo.web.dataadapter.gff3.Gff3DataAdapter</class>

Should point to the class that will handle the write request. Writing is handled by classes that implement the org.bbop.apollo.web.dataadapter.DataAdapter interface. This allows you to add support for writing to different formats. We currently only have support for:

<!-- required permission for using data adapter
 available options are: read, write, publish -->
<permission>publish</permission>

Required user permission for accessing this data adapter. If the user does not have the required permission, it will not be available in the list of data adapters. Available permissions are read, write, and publish.

<!-- configuration for data adapter -->
<config>/config/gff3_config.xml</config>

File that contains the configuration for the data adapter plugin chosen.

<!-- options to be passed to data adapter -->
<options>output=file&amp;format=gzip</options>

Options to be passed to the data adapter. These are dependent on the data adapter.

Next, let’s look at the <data_adapter_group> element:

<!-- display name for adapter group -->
<key>FASTA</key>

This is a string that will be used for the data adapter submenu name.

<permission>read</permission> Required user permission for accessing this data adapter group. If the user does not have the required permission, it will not be available in the list of data adapters. Available permissions are read, write, and publish.

Translation tables

WebApollo has support for alternate translation tables. For your convenience, WebApollo comes packaged with the current NCBI translation tables. They reside in the config/translation_tables directory in your installation (/var/lib/tomcat7/webapps/WebApollo/config/translation_tables). They’re all named ncbi_#_translation_table.txt where # represents the NCBI translation table number (for example, for ciliates, you’d use ncbi_6_translation_table.txt).

You can also customize your own translation table. The format is tab delimited, with each entry containing either 2 or 3 columns. The 3rd column is only used in the cases of start and stop codons. You only need to put entries for codons that differ from the standard translation table (#1). The first column has the codon triplet and the second has the IUPAC single letter representation for the translated amino acid. The stop codon should be represented as * (asterisk).

TAA   Q

As mentioned previously, you’ll only need the 3rd column for start and stop codons. To denote a codon as a start codon, put in start in the third column. For example, if we wanted to assign GTG as a start codon, we’d enter:

GTG   V   start

For stop codons, if we enter an IUPAC single letter representation for the amino acid in the 3rd column, we’re denoting that amino acid to be used in the case of a readthrough stop codon. For example, to use pyrrolysine, we’d enter:

TAG   *   O

If you write your own customized translation table, make sure to update the <translation_table> element in your configuration to your customized file.

Canned comments

You can configure a set of predefined comments that will be available for users when adding comments through a dropdown menu. The configuration is stored in /var/lib/tomcat7/webapps/WebApollo/config/canned_comments.xml. Let’s take a look at the configuration file.

<?xml version="1.0" encoding="UTF-8"?>
 
<canned_comments>
    <!-- one <comment> element per comment.
 it must contain either the attribute "feature_type" that defines
 the type of feature this comment will apply to or the attribute "feature_types"
 that defines a list (comma separated) of types of features this comment will
 apply to.
 types must be be in the form of "CV:term" (e.g., "sequence:gene")
 
 <comment feature_type="sequence:gene">This is a comment for sequence:gene</comment>
 or
 <comment feature_types="sequence:tRNA,sequence:ncRNA">This is a comment for both sequence:tRNA and sequence:ncRNA</comment>
 -->
</canned_comments>

You’ll need one <comment> element for each predefined comment. The element needs to have either a feature_type attribute in the form of CV:term that this comment applies to or a feature_types attribute, a comma separated list of types this comment will apply to, where each type is also in the form of CV:term. Let’s make a few comments for feature of type sequence:gene and sequence:transcript, sequence:mRNA:

<comment feature_type="sequence:gene">This is a comment for a gene</comment>
<comment feature_type="sequence:gene">This is another comment for a gene</comment>
<comment feature_types="sequence:transcript,sequence:mRNA">This is a comment for both a transcript or mRNA</comment>

All supported annotation types can be used.

Search tools

As mentioned previously, WebApollo makes use of tools for sequence searching rather than employing its own search algorithm. The only currently supported tool is command line Blat.

Blat

You’ll need to have Blat installed and a search database with your genomic sequences available to make use of this feature. You can get documentation on the Blat command line suite of tools at BLAT Suite Program Specifications and User Guide and get information on setting up the tool in the official BLAT FAQ. The configuration is stored in /var/lib/tomcat7/webapps/WebApollo/config/blat_config.xml. Let’s take a look at the configuration file:

<?xml version="1.0" encoding="UTF-8"?>
 
<!-- configuration file for setting up command line Blat support -->
 
<blat_config>
 
    <!-- path to Blat binary →
 <blat_bin>ENTER_PATH_TO_BLAT_BINARY</blat_bin>
 
 <!-- path to where to put temporary data -->
    <tmp_dir>ENTER_PATH_FOR_TEMPORARY_DATA</tmp_dir>
 
    <!-- path to Blat database -->
    <database>ENTER_PATH_TO_BLAT_DATABASE</database>
 
    <!-- any Blat options (directly passed to Blat) e.g., -minMatch -->
    <blat_options>ENTER_ANY_BLAT_OPTIONS</blat_options>
 
    <!-- true to remove temporary data path after search (set to false for debugging purposes) -->
    <remove_tmp_dir>true</remove_tmp_dir>
 
</blat_config>

Let’s look at each element with values filled in.

<!-- path to Blat binary -->
<blat_bin>/usr/local/bin/blat</blat_bin>

We need to point to the location where the Blat binary resides. We’ve installed Blat in /usr/local/bin.

<!-- path to where to put temporary data -->
<tmp_dir>/data/webapollo/blat/tmp</tmp_dir>

We need to point to the location where to store temporary files to be used in the Blat search. It can be set to whatever location you’d like.

<!-- path to Blat database -->
<database>/data/webapollo/blat/db/genomic.2bit</database>

We need to point to the location of the search database to be used by Blat. See the Blat documentation for more information on generation search databases.

<!-- any Blat options (directly passed to Blat) e.g., -minMatch -->
<blat_options>-minScore=100 -minIdentity=60</blat_options>

Here we can configure any extra options to used by Blat. These options are passed verbatim to the program. In this example, we’re passing the -minScore parameter with a minimum score of 100 and the -minIdentity parameter with a value of 60 (60% identity). See the Blat documentation for information of all available options.

<!-- true to remove temporary data path after search (set to false for debugging purposes) -->
<remove_tmp_dir>true</remove_tmp_dir>

Whether to delete the temporary files generated for the BLAT search. Set it to false to not delete the files after the search, which is useful for debugging why your search may have failed or returned no results.

Data adapters

GFF3

The GFF3 data adapter will allow exporting the current annotations as a GFF3 file. You can get more information about the GFF3 format at The Sequence Ontology GFF3 page. The configuration is stored in /var/lib/tomcat7/webapps/WebApollo/config/gff3_config.xml. Let’s take a look at the configuration file:

<?xml version="1.0" encoding="UTF-8"?>
 
<!-- configuration file for GFF3 data adapter -->
 
<gff3_config>
 
    <!-- path to where to put generated GFF3 file.  This path is 
     relative path that will be where you deployed your 
     instance (so that it's accessible from HTTP download requests) -->
    <tmp_dir>tmp</tmp_dir>
 
    <!-- value to use in the source column (column 2) of the generated
     GFF3 file. -->
    <source>.</source>
 
    <!-- which metadata to export as an attribute - optional.
     Default is to export everything except owner, date_creation, and date_last_modified -->
    <!--
 <metadata_to_export>
     <metadata type="name" />
     <metadata type="symbol" />
     <metadata type="description" />
     <metadata type="status" />
     <metadata type="dbxrefs" />
     <metadata type="attributes" />
     <metadata type="comments" />
     <metadata type="owner" />
     <metadata type="date_creation" />
     <metadata type="date_last_modified" />
 </metadata_to_export>
 -->
 
    <!-- whether to export underlying genomic sequence - optional.
 Defaults to true -->
    <export_source_genomic_sequence>true</export_source_genomic_sequence>
 
</gff3_config>

<tmp_dir>tmp</tmp_dir>

This is the root directory where the GFF3 files will be generated. The actual GFF3 files will be in subdirectories that are generated to prevent collisions from concurrent requests. This directory is relative to /var/lib/tomcat7/webapps/WebApollo. This is done to allow the generated GFF3 to be accessible from HTTP requests.

<!-- value to use in the source column (column 2) of the generated
 GFF3 file. -->
<source>.</source>

This is what to put as the source (column 2) in the generated GFF3 file. You can change the value to anything you’d like.

<!-- which metadata to export as an attribute - optional.
 Default is to export everything except owner, date_creation, and date_last_modified -->
<metadata_to_export>
    <metadata type="name" />
    <metadata type="symbol" />
    <metadata type="description" />
    <metadata type="status" />
    <metadata type="dbxrefs" />
    <metadata type="attributes" />
    <metadata type="comments" />
    <metadata type="owner" />
    <metadata type="date_creation" />
    <metadata type="date_last_modified" />
</metadata_to_export>

This defines which metadata to export in the GFF3 (in column 9). This configuration is optional. The default is to export everything except owner, date_creation, and date_last_modified. You need to define one <metadata<> element with the appropriate type attribute per metadata type you want to export. Available types are:

<!-- whether to export underlying genomic sequence - optional.
Defaults to true -->
<export_source_genomic_sequence>true</export_source_genomic_sequence>

Determines whether to export the underlying genomic sequence as FASTA attached to the GFF3 file. Set to false to disable it. Defaults to true.

Note that the generated files will reside in that directory indefinitely to allow users to download them. You’ll need to eventually remove those files to prevent the file system from cluttering up. There’s a script that will traverse the directory and remove any files that are older than a provided time and cleanup directories as they become empty. It’s recommended to setup this script as a cron job that runs hourly to remove any files older than an hour (should provide plenty of time for users to download those files). The script is in ~/WebApollo-2014-04-03/tools/cleanup/remove_temporary_files.sh.

$ ~/WebApollo-2014-04-03/tools/cleanup/remove_temporary_files.sh -d /var/lib/tomcat7/webapps/WebApollo/tmp -m 60

Chado

The Chado data adapter will allow writing the current annotations to a Chado database. You can get more information about the Chado at GMOD Chado page. The configuration is stored in /var/lib/tomcat7/webapps/WebApollo/config/chado_config.xml. Let’s take a look at the configuration file:

<?xml version="1.0" encoding="UTF-8"?>
 
<!-- configuration file for Chado data adapter -->
 
<chado_config>
 
    <!-- Hibernate configuration file for accessing Chado database -->
    <hibernate_config>/config/hibernate.xml</hibernate_config>
 
</chado_config>

There’s only one element to be configured:

<hibernate_config>/config/hibernate.xml</hibernate_config>

This points to the Hibernate configuration for accessing the Chado database. Hibernate provides an ORM (Object Relational Mapping) for relational databases. This is used to access the Chado database. The Hibernate configuration is stored in /var/lib/tomcat7/webapps/WebApollo/config/hibernate.xml. It is quite large (as it contains a lot of mapping resources), so let’s take a look at the parts of the configuration file that are of interest (near the top of the file):

<?xml version="1.0" encoding="UTF-8"?>
<!DOCTYPE hibernate-configuration PUBLIC
      "-//Hibernate/Hibernate Configuration DTD 3.0//EN"
      "http://hibernate.sourceforge.net/hibernate-configuration-3.0.dtd">
<hibernate-configuration>
    <session-factory name="SessionFactory">
        <property name="hibernate.connection.driver_class">org.postgresql.Driver</property>
        <property name="hibernate.connection.url">ENTER_DATABASE_CONNECTION_URL</property>
        <property name="hibernate.connection.username">ENTER_USERNAME</property>
        <property name="hibernate.connection.password">ENTER_PASSWORD</property>
 
        ...
 
    </session-factory>
</hibernate-configuration>

Let’s look at each element:

<property name="hibernate.connection.driver_class">org.postgresql.Driver</property>

The database driver for the RDBMS where the Chado database exists. It will most likely be PostgreSQL (as it’s the officially recommended RDBMS for Chado), in which case you should leave this at its default value.

<property name="hibernate.connection.url">jdbc:postgresql://localhost:5432/drupal</property>

JDBC URL to connect to the Chado database. It will be in the form of jdbc:$RDBMS://$SERVERNAME:$PORT/$DATABASE_NAME where $RDBMS is the RDBMS used for the Chado database, $SERVERNAME is the server’s name, $PORT is the database port, and $DATABASE_NAME is the database’s name. Let’s say we’re connecting to a Chado database running on PostgreSQL on server my_server, port 5432 (PostgreSQL’s default), and a database name of my_organism, the connection URL will look as follows: jdbc:postgresql://my_server:5432/my_organism.

<property name="hibernate.connection.username">ubuntu</property>

User name used to connect to the database. This user should have write privileges to the database.

<property name="hibernate.connection.password">none</property>

Password for the provided user name.

FASTA

The FASTA data adapter will allow exporting the current annotations to a FASTA file. The configuration is stored in /var/lib/tomcat7/webapps/WebApollo/config/fasta_config.xml. Let’s take a look at the configuration file:

<?xml version="1.0" encoding="UTF-8"?>
 
<!-- configuration file for FASTA data adapter -->
 
<fasta_config>
 
    <!-- path to where to put generated FASTA file.  This path is a
 relative path that will be where you deployed your WebApollo
 instance (so that it's accessible from HTTP download requests) -->
    <tmp_dir>tmp</tmp_dir>
 
    <!-- feature types to process when dumping FASTA sequence -->
    <feature_types>
 
        <!-- feature type to process - one element per type -->
        <feature_type>sequence:mRNA</feature_type>
 
        <feature_type>sequence:transcript</feature_type>
 
    </feature_types>
 
    <!-- which metadata to export as an attribute - optional.
 Default does not export any metadata -->
    <!--
 <metadata_to_export>
     <metadata type="name" />
     <metadata type="symbol" />
     <metadata type="description" />
     <metadata type="status" />
     <metadata type="dbxrefs" />
     <metadata type="attributes" />
     <metadata type="comments" />
     <metadata type="owner" />
     <metadata type="date_creation" />
     <metadata type="date_last_modified" />
 </metadata_to_export>
 -->
 
</fasta_config>

<!-- path to where to put generated FASTA file.  This path is a
relative path that will be where you deployed your WebApollo
instance (so that it's accessible from HTTP download requests) -->
<tmp_dir>tmp</tmp_dir>

This is the root directory where the FASTA files will be generated. The actual FASTA files will be in subdirectories that are generated to prevent collisions from concurrent requests. This directory is relative to /var/lib/tomcat7/webapps/WebApollo. This is done to allow the generated FASTA to be accessible from HTTP requests.

<!-- feature types to process when dumping FASTA sequence -->
<feature_types>
 
    <!-- feature type to process - one element per type -->
    <feature_type>sequence:mRNA</feature_type>
 
    <feature_type>sequence:transcript</feature_type>
 
</feature_types>

This defines which annotation types should be processed when exporting the FASTA data. You’ll need one <feature_type> element for each type you want to have processed. Only the defined feature_type elements will all be processed, so you might want to have different configuration files for processing different types of annotations (which you can point to in FASTA data adapter in the config element in config.xml). All supported annotation types can be used for the value of feature_type, with the addition of sequence:exon.

In config.xml, in the <options> element in the <data_adapter> configuration for the FASTA adapter, you’ll notice that there’s a seqType option. You can change that value to modify which type of sequence will be exported as FASTA. Available options are:

<!-- which metadata to export as an attribute - optional.
Default does not export any metadata -->
<!--
<metadata_to_export>
 <metadata type="name" />
 <metadata type="symbol" />
 <metadata type="description" />
 <metadata type="status" />
 <metadata type="dbxrefs" />
 <metadata type="attributes" />
 <metadata type="comments" />
 <metadata type="owner" />
 <metadata type="date_creation" />
 <metadata type="date_last_modified" />
</metadata_to_export>
-->

Defines which metadata to export in the defline for each feature. The default is to not output any of the listed metadata. Uncomment to turn on this option. Note that you can remove (or comment) any <metadata> elements that you’re not interested in exporting.

Note that like the GFF3 adapter, the generated files will reside in that directory indefinitely to allow users to download them. You’ll need to eventually remove those files to prevent the file system from cluttering up. You can use the remove_temporary_files.sh script to handle the cleanup. In fact, if you configure both the GFF3 and FASTA adapters to use the same temporary directory, you’ll only need to worry about cleanup from a single location. See the GFF3 section for information about remove_temporary_files.sh.

Data generation

The steps for generating data (in particular static data) are mostly similar to JBrowse data generation steps, with some extra steps required. The scripts for data generation reside in /var/lib/tomcat7/webapps/WebApollo/jbrowse/bin. Let’s go into WebApollo’s JBrowse directory.

$ cd /var/lib/tomcat7/webapps/WebApollo/jbrowse

It will make things easier if we make sure that the scripts in the bin directory are executable.

$ chmod 755 bin/*

As mentioned previously, the data resides in the data directory by default. We can symlink /data/webapollo/jbrowse giving you a lot of flexibility in allowing your WebApollo instance to easily point to a new data directory.

$ ln -sf /data/webapollo/jbrowse data

DNA track setup

The first thing we need to do before processing our evidence is to generate the reference sequence data to be used by JBrowse. We’ll use the prepare-refseqs.pl script.

$ bin/prepare-refseqs.pl --fasta ~/sample_data/pyu_data/scf1117875582023.fa

We now have the DNA track setup. Note that you can also use a GFF3 file containing the genomic sequence by using the --gff option instead of --fasta and point it to the GFF3 file.

Adding the WebApollo plugin

We now need to setup the data configuration to use the WebApollo plugin. We’ll use the add-webapollo-plugin.pl script to do so.

$ bin/add-webapollo-plugin.pl -i data/trackList.json

Static data generation

Generating data from GFF3 works best by having a separate GFF3 per source type. If your GFF3 has all source types in the same file, we need to split up the GFF3. We can use the split_gff_by_source.pl script in ~/WebApollo-2014-04-03/tools/data to do so. We’ll output the split GFF3 to some temporary directory (we’ll use ~/scratch/split_gff).

$ mkdir -p ~/scratch/split_gff
$ ~/WebApollo-2014-04-03/tools/data/split_gff_by_source.pl \
-i ~/sample_data/pyu_data/scf1117875582023.gff -d ~/scratch/split_gff

If we look at the contents of ~/scratch/split_gff, we can see we have the following files:

$ ls ~/scratch/split_gff
blastn.gff  est2genome.gff  protein2genome.gff  repeatrunner.gff
blastx.gff  maker.gff       repeatmasker.gff    snap_masked.gff

We need to process each file and create the appropriate tracks.

GFF3 with gene/transcript/exon/CDS/polypeptide features

We’ll start off with maker.gff. We need to handle that file a bit differently than the rest of the files since the GFF represents the features as gene, transcript, exons, and CDSs.

$ bin/flatfile-to-json.pl --gff ~/scratch/split_gff/maker.gff \
--arrowheadClass trellis-arrowhead --getSubfeatures \
--subfeatureClasses '{"wholeCDS": null, "CDS":"brightgreen-80pct", "UTR": "darkgreen-60pct", "exon":"container-100pct"}' \
--cssClass container-16px --type mRNA --trackLabel maker

Note that brightgreen-80pct, darkgreen-60pct, container-100pct, container-16px, gray-center-20pct are all CSS classes defined in WebApollo stylesheets that describe how to display their respective features and subfeatures. WebApollo also tries to use reasonable default CSS styles, so it is possible to omit these CSS class arguments. For example, to accept default styles for maker.gff, the above could instead be shortened to:

$ bin/flatfile-to-json.pl --gff ~/scratch/split_gff/maker.gff \
--getSubfeatures --type mRNA --trackLabel maker

See the Customizing features section for more information on CSS styles. There are also many other configuration options for flatfile-to-json.pl, see JBrowse data formatting for more information.

GFF3 with match/match_part features

Now we need to process the other remaining GFF3 files. The entries in those are stored as “match/match_part”, so they can all be handled in a similar fashion.

We’ll start off with blastn as an example.

$ bin/flatfile-to-json.pl --gff ~/scratch/split_gff/blastn.gff \
--arrowheadClass webapollo-arrowhead --getSubfeatures \
--subfeatureClasses '{"match_part": "darkblue-80pct"}' \
--cssClass container-10px --trackLabel blastn

Again, container-10px and darkblue-80pct are CSS class names that define how to display those elements. See the Customizing features section for more information.

We need to follow the same steps for the remaining GFF3 files. It can be a bit tedious to do this for the remaining six files, so we can use a simple Bash shell script to help us out (write the script to a file and execute as shown below). Don’t worry if the script doesn’t make sense, you can always process each file manually on the command line:

  for i in $(ls ~/scratch/split_gff/*.gff | grep -v maker); do
    j=$(basename $i)
    j=${j/.gff/}
    echo "Processing $j"
    bin/flatfile-to-json.pl --gff $i --arrowheadClass webapollo-arrowhead \
    --getSubfeatures --subfeatureClasses "{\"match_part\": \"darkblue-80pct\"}" \
    --cssClass container-10px --trackLabel $j
  done

Generate searchable name index

Once data tracks have been created, you will need to generate a searchable index of names using the generate-names.pl script:

$ bin/generate-names.pl

This script creates an index of sequence names and feature names in order to enable auto-completion in the navigation text box. This index is required, so if you do not wish any of the feature tracks to be indexed for auto-completion, you can instead run generate-names.pl immediately after running prepare_refseqs.pl, but before generating other tracks.

The script can be also rerun after any additional tracks are generated if you wish feature names from that track to be added to the index (using the --incremental option).

BAM data

Now let’s look how to configure BAM support. WebApollo has native support for BAM, so no extra processing of the data is required.

First we’ll copy the BAM data into the WebApollo data directory. We’ll put it in the data/bam directory. Keep in mind that this BAM data was randomly generated, so there’s really no biological meaning to it. We only created it to show BAM support.

$ mkdir data/bam
$ cp ~/sample_data/pyu_data/*.bam* data/bam

Now we need to add the BAM track.

$ bin/add-bam-track.pl --bam_url bam/simulated-sorted.bam \
   --label simulated_bam --key "simulated BAM"

You should now have a simulated BAM track available.

BigWig data

WebApollo has native support for BigWig files (.bw), so no extra processing of the data is required.

Configuring a BigWig track is very similar to configuring a BAM track. First we’ll copy the BigWig data into the WebApollo data directory. We’ll put it in the data/bigwig directory. Keep in mind that this BigWig data was generated as a coverage map derived from the randomly generated BAM data, so like the BAM data there’s really no biological meaning to it. We only created it to show BigWig support.

$ mkdir data/bigwig
$ cp ~/sample_data/pyu_data/*.bw data/bigwig

Now we need to add the BigWig track.

$ bin/add-bw-track.pl --bw_url bigwig/simulated-sorted.coverage.bw \
  --label simulated_bw --key "simulated BigWig"

You should now have a simulated BigWig track available.

Customizing different annotation types

To change how the different annotation types look in the annotation track, you’ll need to update the mapping of the annotation type to the appropriate CSS class. This data resides in trackList.json after running add-webapollo-plugin.pl. You’ll need to modify the JSON entry whose label is Annotations. Of particular interest is the alternateClasses element. Let’s look at that default element:

"alternateClasses": {
    "pseudogene" : {
       "className" : "light-purple-80pct",
       "renderClassName" : "gray-center-30pct"
    },
    "tRNA" : {
       "className" : "brightgreen-80pct",
       "renderClassName" : "gray-center-30pct"
    },
    "snRNA" : {
       "className" : "brightgreen-80pct",
       "renderClassName" : "gray-center-30pct"
    },
    "snoRNA" : {
       "className" : "brightgreen-80pct",
       "renderClassName" : "gray-center-30pct"
    },
    "ncRNA" : {
       "className" : "brightgreen-80pct",
       "renderClassName" : "gray-center-30pct"
    },
    "miRNA" : {
       "className" : "brightgreen-80pct",
       "renderClassName" : "gray-center-30pct"
    },
    "rRNA" : {
       "className" : "brightgreen-80pct",
       "renderClassName" : "gray-center-30pct"
    },
    "repeat_region" : {
       "className" : "magenta-80pct"
    },
    "transposable_element" : {
       "className" : "blue-ibeam",
       "renderClassName" : "blue-ibeam-render"
    }
},

For each annotation type, you can override the default class mapping for both className and renderClassName to use another CSS class. Check out the Customizing features section for more information on customizing the CSS classes.

Customizing features

The visual appearance of biological features in WebApollo (and JBrowse) is handled by CSS stylesheets. Every feature and subfeature is given a default CSS “class” that matches a default CSS style in a CSS stylesheet. These styles are are defined in /var/lib/tomcat7/webapps/WebApollo/jbrowse/track_styles.css and /var/lib/tomcat7/webapps/WebApollo/jbrowse/plugins/WebApollo/css/webapollo_track_styles.css. Additional styles are also defined in these files, and can be used by explicitly specifying them in the -cssClass, –subfeatureClasses, –renderClassname, or –arrowheadClass parameters to flatfile-to-json.pl. See example above

WebApollo differs from JBrowse in some of it’s styling, largely in order to help with feature selection, edge-matching, and dragging. WebApollo by default uses invisible container elements (with style class names like “container-16px”) for features that have children, so that the children are fully contained within the parent feature. This is paired with another styled element that gets rendered within the feature but underneath the subfeatures, and is specified by the –renderClassname argument to flatfile-to-json.pl. Exons are also by default treated as special invisible containers, which hold styled elements for UTRs and CDS.

It is relatively easy to add other stylesheets that have custom style classes that can be used as parameters to flatfile-to-json.pl. Examples are available in /var/lib/tomcat7/webapps/WebApollo/jbrowse/plugins/WebApollo/css/custom_track_styles.css.

You need to tell WebApollo where to find these styles. Additional CSS files can be specified in the trackList.json file that is created in the data directory during static data generation, by adding a “css” property to the JSON data:

   "css" : "plugins/WebApollo/css/custom_track_styles.css"

Then these new styles can be used as arguments to flatfile-to-json.pl, for example:

bin/flatfile-to-json.pl --gff /split_gff/maker.gff 
--getSubfeatures --type mRNA --trackLabel maker --webApollo 
--subfeatureClasses '{"CDS":"darkorange-80pct", "UTR": "darkorange-60pct"}'

Restart WebApollo

It is generally a good idea to restart WebApollo (or Tomcat) after making any configuration modifications. Note that you don’t need to restart the application if you’ve only modified data.

$ sudo service tomcat7 restart

Bulk loading annotations to the user annotation track

GFF3

You can use the ~/WebApollo-2014-04-03/tools/data/add_transcripts_from_gff3_to_annotations.pl script to bulk load GFF3 files with transcripts to the user annotation track. Let’s say we want to load our maker.gff transcripts.

$ ~/WebApollo-2014-04-03/tools/data/add_transcripts_from_gff3_to_annotations.pl \
-U localhost/WebApollo -u web_apollo_admin -p web_apollo_admin \
-i ~/scratch/split_gff/maker.gff

The default options should be handle GFF3 most files that contain genes, transcripts, and exons.

You can still use this script even if the file you’re loading does not contain transcripts and exons. Let’s say we want to load match and match_part features as transcripts and exons respectively. We’ll use the blastn.gff file as an example.

$ ~/WebApollo-2014-04-03/tools/data/add_transcripts_from_gff3_to_annotations.pl \
-U localhost/WebApollo -u web_apollo_admin -p web_apollo_admin \
-i ~/scratch/split_gff/blastn.gff -t match -e match_part

Look at the script’s help (-h) for all available options.

Testing out your installation

Let’s test out our installation. Point your browser to http://ec2-##-##-##-##.compute-1.amazonaws.com/WebApollo
(e.g., http://ec2-184-73-92-243.compute-1.amazonaws.com/WebApollo).

WebApollo login page

The user name and password are both web_apollo_admin as we configured earlier.

WebApollo reference sequence selection

We only see one reference sequence to annotate since we’re only working with one contig. Click on the ` scf1117875582023` link.

Congratulations, you’re done configuring WebApollo!

Now have fun annotating!!!

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