Question: Which package to use for RPKM/FPKM RNA-seq counts?
0
gravatar for hafiz.talhamalik
4 weeks ago by
Pakistan
hafiz.talhamalik0 wrote:

I have RNA-Seq counts which are FPKM/RPKM normalized. which package in R should i use for Differential gene expression. among many packages which package is better? is DGE using raw count is better or using FPKM/RPKM normalized counst are better ?

Thanks in anticipation.

rna-seq • 172 views
ADD COMMENTlink modified 4 weeks ago by Renesh1.1k • written 4 weeks ago by hafiz.talhamalik0
2
gravatar for Macspider
4 weeks ago by
Macspider1.5k
Vienna - BOKU
Macspider1.5k wrote:

which package in R should i use for Differential gene expression?

Assuming you mean to do diff.expr. with FPKM, I think Limma does a pretty good job. However ...

among many packages which package is better?

This question I think is hard to answer. The jury is out on this one!

is DGE using raw count is better or using FPKM/RPKM normalized counst are better ?

The author of the FPKM measurements himself called FPKM as "biased" and "wrong". So I would go for raw counts when possible. My pipeline usually includes htseq-count and DESeq2, but other couting programs and other differential expression analysis programs are fine!

ADD COMMENTlink written 4 weeks ago by Macspider1.5k

Thanks.. but limma was built for micro-array is it good enough for RNA-seq ?

ADD REPLYlink written 4 weeks ago by hafiz.talhamalik0

As far as I know, you can plug FPKM values in.

ADD REPLYlink written 4 weeks ago by Macspider1.5k
0
gravatar for Renesh
4 weeks ago by
Renesh1.1k
United States
Renesh1.1k wrote:

DESeq2 is a good package for DEG analysis in R. https://bioconductor.org/packages/release/bioc/html/DESeq2.html

If you are using R package for DEG analysis, then go ahead with raw counts. It is not recommended to perform DEG analysis on normalized count using DESeq2 because it performs internal normalization.

ADD COMMENTlink written 4 weeks ago by Renesh1.1k

Thankyou so much.. and what about edgeR ?

ADD REPLYlink written 4 weeks ago by hafiz.talhamalik0
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