data input for GSEA: TPM or R/FPKM vs. normalized counts

[Dong]

Hi Anthony, 

I just notice you ever stated that neither TPM or R/FPKM are considered the appropriate normalization to use for computing differential expression between samples.  Would you be able to provide the reason for this?  That will be great if you can also provide some references. From the video delivered by QUEST, I thought TPM would be more appropriate for computing differential expression between samples. 

Thanks.

Dong

[Anthony Castanza]

Hi Dong,

I don't know anything about the QUEST video, but this primer is a pretty good explanation of why they aren't appropriate: https://hbctraining.github.io/DGE_workshop/lessons/02_DGE_count_normalization.html

You can also see a response from Gordon Smyth (the PI of the lab that developed the the limma-voom pipeline) here: https://support.bioconductor.org/p/98820/#98875 specifically advising against the use of TPM for differential expression (of which the same concerns apply to FPKM)

Hope that helps!

-Anthony

Anthony S. Castanza, PhD

Curator, Molecular Signatures Database

Mesirov Lab, Department of Medicine

University of California, San Diego

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[Dong]

Hi Anthony, 

Thanks for sharing the information.  They are really important and helpful.

Best,

Dong