Question: best workflow for RNA data analysis without replications
4
gravatar for Chao.wang2
3.6 years ago by
Chao.wang240
Canada
Chao.wang240 wrote:

I am wondering could anyone tell me the best work flow (software) and Statistical model should I use for RNA seq data (without replications).

 

 

Thanks a lot

rna-seq • 2.8k views
ADD COMMENTlink modified 3.6 years ago by Devon Ryan91k • written 3.6 years ago by Chao.wang240
3

What are you trying to do with the RNA-seq data?  If you have no replicates, you should not attempt to assess differential expression.

ADD REPLYlink written 3.6 years ago by Rob3.4k

Thanks for your reply, actually, I want to assess the differential expression. If I can't assess differential exprssion, what else can I do with this kind of data.

Thanks

ADD REPLYlink written 3.6 years ago by Chao.wang240

Not much, you can't really do much of the way of comparisons. You could ask whether a gene is likely expressed, or what its major isoform is, but even then any result would have the "at least in this sample" caveat applied to it.
 

ADD REPLYlink written 3.6 years ago by Devon Ryan91k
1

edgeR authors provide several solutions for people who work without replicates: https://bioconductor.org/packages/release/bioc/vignettes/edgeR/inst/doc/edgeRUsersGuide.pdf (page 21). However, I agree with previous comment that you should avoid working without replicates. 

ADD REPLYlink written 3.6 years ago by mkulecka300

thank you for your advice.

ADD REPLYlink written 3.6 years ago by Chao.wang240

You can analyse this data by fold change between the groups. But the data should be normalized. Is not the best way to avaluate rnaseq, but as you dont have replicates you can validate some of most down or up regulated rnas in your data by Real Time PCR, if is possible.

ADD REPLYlink written 3.6 years ago by silas008100
2
gravatar for Devon Ryan
3.6 years ago by
Devon Ryan91k
Freiburg, Germany
Devon Ryan91k wrote:

The best tool you'll find is GFold, and you should obviously be very cognizant of the caveats that come with trying to draw any firm conclusions from unreplicated data (i.e., the most this is useful for is getting some target genes for replicated targeted tests).

ADD COMMENTlink written 3.6 years ago by Devon Ryan91k

Thanks, I will check that.

ADD REPLYlink written 3.6 years ago by Chao.wang240
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