24 April 2014

BLAST

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Thursday notes

ApTranscriptome

As a complement to topGO approach, use DAVID to focus on genes with high-quality annotations in model organisms Apis and Drosophila.

Get these BLAST hits using method for annotation I tried before going to FastAnnotator.

• Found Taxids on NCBI for
• Apis Taxid: 7459
• Drosophila Taxid: 7215
• Searched Entrez Protein Database with query txid7215[ORGN] OR txid7459[ORGN]
• Selected “Send to File” wiht format “GI list”
  • generated list of 585,719 unique gi ids

• Used list of GIs from the previous step with the blastdb_aliastool to build an aliased blastdb of just insects:

  blastdb_aliastool -gilist apis_drosophila.gi.txt -db nr -out nr_apis_drosophila -title “Apis and Drosophila nr database”

which gave output:

Converted 585719 GIs from apis_drosophila.gi.txt to binary format in nr_apis_drosophila.p.gil Created protein nr_apis_drosophila BLAST (alias) database with 274099 sequences (out of 585719 in nr_apis_drosophila.p.gil, 47% found)

• Tested search against new (aliased) database:

  blastx -query query.fa -db nr_insecta -out querytest

Worked!

Now…for the blastx of the whole caboodle, given problems I had before, trialed and used GNU parallel as described on Biostars to efficiently use all cores.

Test script for 50 sequences

cat ../test.fa | parallel --block 100k --recstart '>' --pipe blastx -evalue 0.01 -outfmt 6 -db /home/data/databases/ncbi_db/nr_apis_drosophila -query - > result

Moved analysis to Mason cluster - already had NCBI nr database downloaded there. Started script running…

CCThermBe

Meet with Grace to go over results.

Ideas for continuation of Grace’s project:

1. Positive control - use juvenile hormone which is known to promote foraging (Page et al. unknown)

• juvenile hormone analog methoprene used by (Cahan et al. 2011)

2. If this works (e.g. treated callows more likely to become foragers), test suite of other environmental factors:

• paraquat
• infection
• development-heat stress
• etc.

3. Otherwise, initiate new experiment:

• thermal adaptation of Aphaenogaster along elevational gradients in VT
• gene expression response to fungal infection

Bibliography

• Sara Helms Cahan, Christopher J. Graves, Colin S. Brent, (2011) Intergenerational Effect of Juvenile Hormone on Offspring in Pogonomyrmex Harvester Ants. Journal of Comparative Physiology B 181 991-999 10.1007/s00360-011-0587-x
• Robert E. Page, Ricarda Scheiner, Joachim Erber, Gro V. Amdam, (unknown) Current Topics in Developmental Biology. Unknown