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fibar 12-02-2015 10:20 AM

What's the best way to identify the taxonomy of assembled genomes?
Hi everyone,
Briefly, I applied coverage-based binning method plus pentamer-frequency ordination on metagenomic assembled contigs, to isolate individual genomes.

Now I'm looking for clever (confident, efficient) methods to answer following questions:
1) Who do those genomes belong to? (ie. identify their taxonomy)
2) How complete are they?

For (1) I've tried AmphoraNet and found it quite good. But what about blasting everything against NCBI-genomes database? Would that work fast enough on a small server (Xeon, 16 cores, 96 GB memory)?

Question (2) could probably be answered if (1) is reached succesfully.

However, it could happen that I get, e.g., a novel Proteobacteria, but how to be sure this is really the case and it is not due to misassembly issues. It's a challenging discipline.

Thanks for your insights.

Markiyan 03-21-2016 04:03 AM

For the course level use blastx(p), for more refined - blastn
I may suggest trying blastx or blastp (if you've got good protein predictions) for the coarse level of closest neighbour finding.

For more refined use blastn on the DNA sequence (nt or similar db).
It is a good idea to create more specific database (esp nt one), and have them on dedicated SSD (esp. nvme or PCIe one).

On the performance side of things, give blastn 900bp or 1kb chunks, turn megablast on and upp the word size. "-n T -W 32".
For blastx - raise the expect to 1e-5 or 1e-10, and give it 450 bp chunks.

Also it is good idea to play with number of cpus per job (depending on number of cores per cpu, db/ram size).

PS: also there is a blast-like tool called diamond (used by AntiSMASH), which may be a bit faster in some cases, but the developers kept changing it's output format, so not as stable as blast, if you need to write your own parser.

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