Question: RNA-Seq requirements for both gene expression quantification and variant calling
0
gravatar for Ozzie
2.7 years ago by
Ozzie10
Croatia/Zagreb
Ozzie10 wrote:

I would like to use RNA-Seq on fresh tumor tissue for both gene expression quantification and variant calling.

Do I need stranded RNA information?

Do I need polyA enrichment based or rRNA depletion based rRNA removal?

How many data (sequencing reads or coverage) per sample do I need?

Thanx!

ADD COMMENTlink modified 2.7 years ago by Devon Ryan95k • written 2.7 years ago by Ozzie10

Being curious, would like to know the basis of your sample size.

ADD REPLYlink modified 2.7 years ago • written 2.7 years ago by cpad011213k
1
gravatar for Devon Ryan
2.7 years ago by
Devon Ryan95k
Freiburg, Germany
Devon Ryan95k wrote:
  1. No, but it's not much more expensive and you get a bit nicer data.
  2. You can do either, but poly-A enrichment would probably make the most sense.
  3. As much as you can afford if you want to do variant calling. Honestly, I'd do 15-20 million reads for RNAseq and then pair that with exome sequencing. Otherwise you're going to have to really up your depth to get decent coverage of lowly-expressed genes and will still be left with the problem of allele-specific expression.
ADD COMMENTlink written 2.7 years ago by Devon Ryan95k
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