We designed an experiment to explore the potential role of carbon dioxide on algae physiology using RNA-Seq. We analyse the differential gene expression using DESeq2 but now we are interested into analyse a few genes in detail.
My PI say to make the statistics over the ration response (expression_treatment/expression_control - 1) and my question is the following: Can I use TPM to calculate this ratio and make following statistics? I read in the documentation of DESeq2 that TPM we can't use this normalisation method to differential expression and my my other question is Why shouldn't?
My advisor ask to re-do the statistical analysis using the ratio response (rr) so i need the rr for each sample. I think this doesnt make sense cause the result will be or should be the same that using DESeq2.
Update: I'm using the fold-change calculated by DESeq2 and I'm not sure if I have transformed the data correctly. I took the two columns
lfcSE (both are log-transformed variables) so I went back to the non-transformed variable as follow:
SE=lfcSE*2^log2FoldChange. I saw this approximation in this post. Is that correct? I have obtained a very broad range of error/