We have identified vaccine candidates via an ex-vivo RNA-seq approach. Next step would be to perform conservation of these candidates (about 20) between multiple bacterial strains (about 200). I would like to know your suggestions to perform this step.

From my bioinformatic experience, it could be possible to start with a first rough screening using blastp or blastx against a multiple strain database, and more accurate approaches would be then performed to validate this results.

Please let me know your suggestions and possible references you might consider helpful to answer this question.

Thanks for your help. Best regards, Bernardo

  • $\begingroup$ It may help to specify how sophisticated your computational techniques are. The answer will be very different depending on whether you want to use websites and black-box software, or if you are able to do this on your own computer using custom-made scripts/programs. $\endgroup$ – adam.r Dec 1 '13 at 18:56
  • $\begingroup$ Hi adam, I prefer command-line tools, I have medium programming experience. I would like to combine already available tools with possible on-the-go scripts if needed. $\endgroup$ – biotech Dec 4 '13 at 17:00

This website will allow you to blast all sequenced microbial genomes: http://www.ncbi.nlm.nih.gov/sutils/genom_table.cgi

Recovering the full gene sequence might be a bit of a hassle, since BLAST will only allow you to recover the region that matched your query. You may need to set up your own pipeline for this. You may also be able to identify a "protein family" or "cluster of orthologous genes" using the BLAST results.

Once you have the full sequence, you can perform a multiple sequence alignment with CLUSTAL, Muscle, or another program. This will help you to identify the conserved regions.

  • $\begingroup$ Hi adam. The 200 genomes have not been published yet, so I would have to use local BLAST in any case. I will avoid to recover full gene sequence going through BLASTP. Thanks for the CLUSTAL, Muscle tools, they will be useful for a latter step. I think the question now is to think how to get yes/no table, that is, presence/absence of each vaccine candidate in each of the 200 bacterial strains. $\endgroup$ – biotech Dec 4 '13 at 17:08

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