[BioC] need help for the study design of a RNA-Seq project

[Sean Davis]

On Thu, Apr 11, 2013 at 4:32 PM, shirley zhang <shirley0818 at gmail.com> wrote:
> Dear list,
>
> I am not sure whether this list is the right place to ask this study design
> question. But on this list, I got lots of information regarding how to
> analyze RNA-Seq data, so would like to give a try.
>
> We are going to do RNA-Sequencing using Illumina HiSeq for 200 samples.
> Given that the sample size is fixed, and the budget is fixed, the following
> 3 options were proposed.
>
> 1.  50bp pair-end reads, sequencing each sample per lane --> we will get
> ~100 million reads per sample
> 2.  75bp pair-end reads, sequencing two samples per lane --> we will get
> ~50-60 million reads per sample
> 3.  100bp pair-end reads, sequencing four samples per lane --> we will get
> ~30-40 million reads per sample
>
> Based on your experience, which option is the best or you have other
> suggestions? We would like to do different kinds of analysis for these
> data, i.e.,novel transcripts, lncRNA, and splicing, SNP, etc. You name it.
> If we have to sort them by priority (from high to low), I would like to say
> " novel transcripts, long-noncoding RNAs splicing and differential
> expression".
>
>  Currently, the majority of labs sequence 100bp pair-end, right? But I was
> told that even you sequence 100bp long, after 75bp, the sequencing quality
> is very bad due to the issue of sequencer itself, that is, it has nothing
> with the RNA quality of samples. If this is true, why is 100bp read length
> becoming more popular now?

Hi, Shirley.

I don't mean this as MY answer to your question, but this blog post
has a few statements that might be interesting to you.

http://core-genomics.blogspot.com/2013/04/encodes-rna-seq-recommendations-need.html

You'll not that it refers to the ENCODE RNA-seq guidelines which might
also be instructive.

Sean