Web Apollo Resources at the National Agricultural Library

Christopher ChildersNAL ARS USDA

i5k.nal.usda.gov

Agenda

• Web Apollo– Web based tool for manual gene curation– Registration required for access

• Introduction to the Web Apollo interface– Viewing evidence tracks– Logging in/out

• BLAST• Example annotation

Annotation Overview

1. Find sequence2. Align to genome3. Examine available evidence4. Annotate based on evidence5. Compare results to related sequences

– BLAST/BLAT or multiple alignment

6. Repeat steps 1 through 5 as needed to refine model7. Add annotation details in the “Information Editor”

Annotator Registration

Some conventions

• HSP – High scoring pair in BLAST/BLAT alignments– The ‘Hits’ in an alignment result set– A subsection of a pair of sequences with sufficient score– Change based on the alignment parameters

• Five prime end and three prime end – Based on direction of transcription– Initiation site is at the five prime end– Stop codon is at the three prime end

• In the genome browser, arrowheads indicate direction3’ 5’

5’ 3’

Web Apollo/JBrowse

• JBrowse is a web based genome browser– Visualize features that are mapped to a genome– These features are displayed as tracks– Many different types of data may be

• Web Apollo adds editing functions to JBrowse– Manual gene curation– Changes automatically saved back to server– Edits are visible to other annotators in real-time

Web Apollo

1

3

2

Viewing evidence

1. The “Available Tracks” panel lists all tracks– Tracks are grouped hierarchically

• Study or analysis• Categories based on evidence type

2. The Navigation panel describes the viewed region– Coordinate ranges for the entire sequence on top– Navigation controls including pan, zoom and search in the middle– Coordinate ranges for the current view along the bottom

3. The Evidence panel shows the features in genomic context– Each type of data is a separate track– Tracks may be displayed in different ways

Viewing evidence

• Turning tracks on or off– Click a tracks checkbox to turn it on or off– Click ‘X’ by the track label also turns the track off

Login

• For scaffold selection view:

Login

• Directly from JBrowse genome browser

Login

• Directly from JBrowse genome browser

Login

• Directly from JBrowse genome browser

BLAST at the i5k Workspace

• Redesigned: uses the latest version of BLAST+• Interface updates options automatically– Based on the combination of query and database– Chooses correct options (blastn, blastx, tblastn, etc)

• Results persist for one week– The results URL can be saved or shared

• HSPs are viewable in the genome browser– HSPs are conservatively chained together– Visible as connected blocks

BLAST

BLAST Results

BLAST results in Web Apollo

Live Annotation Example

• Alpha-catenin in the Colorado Potato Beetle• We created special training versions of our toolsBLAST: https://i5k.nal.usda.gov/training/blast Web Apollo:https://apollo.nal.usda.gov/lepdec_training/jbrowse

Annotation using real data

• Logging in to the demo instance• Possible gene families to look at– Cytochrome P450– myotubularin-related proteins– COP9

Sequence Search

• Map a gene of interest to a genomic region– BLAT: Web Apollo plugin– BLAST@NAL: separate tool

• Each has unique features– BLAT shows the hit as a highlighted region– BLAST shows hits as segments, similar to other

evidence tracks