Hello, my name is Jonathan I'm new to R and to RNA-seq. I have a question regarding a project I'm working on. I want to compare differential expression of RNA within 4 different groups of cells (different tumors) each has about 35 people. I need to look for deferential expression of about 70,000 RNA. can I use DESEQ2 for that even though the cell are not paired or is there any other tool in order to do so? if someone can link me a manual for that kind of analysis I would be grateful. thank you for your support, Jonathan.
If I understand correctly, you have four different tumors, but no reference healthy sample to compare to. In that case, you can take two different approaches with DESeq2 (given that they are comparable/from the same experimental batch, as ATpoint pointed out):
- Either make pairwise comparison of the different groups then combine the results (i.e, genes differentially expressed in tumor A vs B & A vs C & A vs D).
Another possibility is to compare tumor A vs the average of B,C and D. This can be specified with:
results(dds, contrast=list("tumorA", c("tumorB","tumorC","tumorD")), listValues=c(1, -1/3))