Hello,
I have a question regarding edgeR normalization,
I have a three tissue RNAseq dataset, with males and females sequenced for both sexes,
After normalization (TMM for read-depth and "composition"+FPKM for gene length) I got similar median of expression for both sexes for two tissues but not the third one (see below),
Because the third tissue possess a lot of genes that are uniquely expressed in one sex, I tried to perform the normalization utilizing only the genes that are expressed in both sexes of all tissues as control genes (sort of 'housekeeping genes' - see vertical line above for the threshold between expressed and non expressed genes),
However, even this way, the difference persists (see below),
Can someone help me to understand what is going on ?
Thank you, Vincent
New Nature paper: Makes and females are physiologically not the same. Joke aside, which tissue is number 3? A better diagnostic plot would be an MA-plot by the way and not using FPKM since this is not compatible with edgeR (or any) serious testing framework.