Assembly of amplicons

Question

[Picasa]

Hi,

I have a pcr amplicon data sequencing of 5 species and 2 genes markers.

My reads were sequenced with HiSeq; so I got 2 x PE of 125 bp.

After sorting my reads per individual, I would like now to assemble my reads in order to retrieve the 2 genes marker (length of 1000 bp aproximately)

1) Anybody have already done that kind of stuff ? Do you know a specific assembler for this job ?

2) The coverage should be different (due to pcr amplication), iam looking for an assembly who can give me the coverage for the contig.

Thanks for your help

[Dattatray Mongad]

Try doing following steps: 1. use PEAR to merge forward and reverse reads. 2. Then use any denovo assembler to assemble reads. (If u know reference it would be nice) 3. Try to map to reference to validate (if neccessary).

Answer 1

[Brice Sarver]

Try one of these:

1. De novo assemble all of your reads. If your amplicons are in high coverage, you ought to be able to recover them. Any standard assembler will do (ABySS, Velvet, Newblet, SOAPdenovo, etc.)

2. Since you know what you are looking for, you can map your reads to those targets. Extract the reads from the BAM file and assemble them de novo. Have a look at ARC.

[Picasa]

Thanks Brice.

I've never head about ARC.

Did you have any experience on that ? in which way could it be "better" than the standard ones.

[Brice Sarver]

I've used it for a few things, and it's great. I'm one of the authors on the manuscript :)

What do you mean by "standard ones?" Part of the motivation for developing it was that there weren't any approaches that did exactly what we needed.