Dear all,

There seems to be something wrong with my designs because even though I have 4 different variables in my design I will end up with 4 times the same heatmap showing the top differentially expressed genes...

Example:

Formula 1:

dds1 <- DESeqDataSetFromMatrix(countData = cts, colData = colData, design = ~ batch + Gender)

Heatmap: Gene A, Gene B, Gene C...

Formula 2:

dds2 <- DESeqDataSetFromMatrix(countData = cts, colData = colData, design = ~ batch + location)

Heatmap: Same as above. .......

Still the top 20 differentially expressed genes are the same... Or are the gender genes so dominant that they would always come up as the first 20?

I am confused right now, any idea much appreciated :)

Thank you, Bine

It's not clear from above if you are running DESeq separately on each one and then results separately, and then using the separate output of results to make distinct heatmaps.

Forget about the heatmaps, can you confirm that the results tables from distinct dds are identical or not?