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# mRNAseq and SNPs call, how can we make this as good as possible?

Member Posts: 39
edited October 2013

Hi!
I have worked some time on a mRNAseq set, single-end. Its a high quality set and lots of biological replicates (200+).

My question is, how could I best contribute to the methodology used for SNPs call in mRNAseq? What do we need tested to improve this method?

Tagged:

• Dev Posts: 50

Hi,
I'm working on testing and creating a best practices pipeline for RNAseq data and I would be happy to hear and learn what tools and protocol do you use. What do you consider to be hight quality set and how do you evaluate your results. We can discuss it here in the forum or in email, whatever you prefer.

Thanks,
Ami

• Member Posts: 39

Hi!
I have just started the testing and this is my first meeting with NGS, so much is new to me.

So far I have:
Tophat 2 -> different pre. pros. w/ Picard -> GATK "best practise" or Samtools -> Total called SNPs evaluation against each other, dpSNP and 1000G .

Im planning on doing the same with BWA. Also I want to tweak the settings of Haplotype caller more. My computer setup is kinda slow, so doing a lot of test takes time. Thats why I was asking here, so I don't waste time doing something obvious.

I consider my set high quality after FastQC evaluation: 50mill. depth, 51bp length, systematic GC content typical of Illumina 2500, no quality drop from 1-51bp, no Kmer problems. Duplication levels are typical of mRNAseq, but how to handle that is still an issue.

Thanks!