Annotate-a-genome: Difference between revisions
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imported>Weigang m (→Protocol) |
imported>Weigang m (→Protocol) |
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=Protocol= | =Protocol= | ||
==Fetch genome sequences== | |||
<syntaxhighlight lang="bash" line enclose="div"> | <syntaxhighlight lang="bash" line enclose="div"> | ||
./bioseq -z 'b31_accession' -o 'genbank' > b31.gb # Reference genome for ortholog identification. Choose main, cp26, or lp54 | ./bioseq -z 'b31_accession' -o 'genbank' > b31.gb # Reference genome for ortholog identification. Choose main, cp26, or lp54 | ||
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./bioseq -t new.nuc > new.pep | ./bioseq -t new.nuc > new.pep | ||
</syntaxhighlight> | </syntaxhighlight> | ||
==Predict orthologs with reciprocal BLAST== | |||
## Prepare BLAST | <syntaxhighlight lang="bash" line start="7" enclose="div"> | ||
# | makeblastdb -in b31.pep -parse_seqids # Prepare the reference DB | ||
# | makeblastdb -in new.pep -parse_seqids # Prepare the new genome DB | ||
blastp -query new.pep -db b31.pep -outfmt 6 -evalue 1e-3 out forward_blast.out # Forward BLAST | |||
blastp -query b31.pep -db new.pep -outfmt 6 -evalue 1e-3 out reverse_blast.out # Reverse BLAST | |||
./check-reciprocal.pl forward_blast.out reverse_blast.out > new.orthlogs 2> new.not-orthologs # Identify orthologs | |||
</syntaxhighlight> | |||
==Verification with synteny broswer= | |||
<syntaxhighlight lang="bash" line start="12" enclose="div"> | |||
./gb2fas -t new.gb > new-to-orf-table.txt | |||
</syntaxhighlight> | |||
Revision as of 22:08, 13 February 2014
Project Goals
- Annotate and add newly sequenced Borrelia genomes to BorreliaBase
- Build an informatics pipeline for gene prediction, ortholog calls, databasing, and synteny analysis
Download genome sequences from GenBank
| Strain | Species | Group | Genome Sequences | Notes |
|---|---|---|---|---|
| B31 | B. burgdorferi (reference genome) | Lyme Disease | Example | |
| CA382 | B. burgdorferi (California) | Lyme Disease | Example | |
| CA8 | B. burgdorferi (California) | Lyme Disease | Example | |
| BgVir | B. garinii (Russia) | Lyme Disease | Example | |
| NMJW1 | B. garinii (China) | Lyme Disease | Example | |
| HLJ01 | B. afzelii (China) | Lyme Disease | Example | |
| Ly | B. duttonii (Tanzania) | Relapsing Fever |
|
Example |
| A1 | B. recurrentis (Ethiopia) | Relapsing Fever | Example | |
| DAH | B. hermsii (Washington State) | Relapsing Fever | Example | |
| 91E135 | B. turicatae (Texas) | Relapsing Fever | Example | |
| Achema | B. crocidurae (Mauritania) | Relapsing Fever |
|
Example |
| HR1 | B. parkeri (??) | Relapsing Fever | Example | |
| LB-2001 | B. miyamotoi (Northeast US) | Relapsing Fever | Example |
Protocol
Fetch genome sequences
./bioseq -z 'b31_accession' -o 'genbank' > b31.gb # Reference genome for ortholog identification. Choose main, cp26, or lp54
./bioseq -z 'gb_accession' -o 'genbank' > new.gb
./gb2fas -n b31.gb > b31.nuc # Extract CDS
./gb2fas -n new.gb > new.nuc
./bioseq -t b31.nuc > b31.pep # Translate (and remove those with internal stop codons)
./bioseq -t new.nuc > new.pep
Predict orthologs with reciprocal BLAST
makeblastdb -in b31.pep -parse_seqids # Prepare the reference DB
makeblastdb -in new.pep -parse_seqids # Prepare the new genome DB
blastp -query new.pep -db b31.pep -outfmt 6 -evalue 1e-3 out forward_blast.out # Forward BLAST
blastp -query b31.pep -db new.pep -outfmt 6 -evalue 1e-3 out reverse_blast.out # Reverse BLAST
./check-reciprocal.pl forward_blast.out reverse_blast.out > new.orthlogs 2> new.not-orthologs # Identify orthologs
=Verification with synteny broswer
./gb2fas -t new.gb > new-to-orf-table.txt