I have 4 conditions and I want to find up/down regulated genes using DESeq2 by comparing
conditions3. I tried two methods:
- I uploaded all conditions with all 3 replicates and created one single table with normalised raw counts. I used
contrastin order to get the comparison between the desired conditions:
dds <- DESeq(dds) res <- results(dds) res <- results(dds, contrast=c("condition","condition1","condition2")) resultsNames(dds) contrast=c("condition","condition1","condition2") resLFC <- lfcShrink(dds, alpha=0.001, lfcThreshold = 1, contrast=contrast, type="ashr") resLFC up <- subset(resLFC, log2FoldChange > 1) down <- subset(resLFC, log2FoldChange < -1)
- I uploaded raw counts only for
condition2, normalised and performed DESeq2 and used the same
My question is, why am I getting different results (i.e. number of up/down regulated genes)? Which is the right/best way? Finally, how should I treat the biological replicates (merge them (if yes,when?) or keep them as separate samples).