Hi Michael and community,
My question this time is about how to set up the Wald test log2 fold change above a threshold, and how to interpret the result correctly.
Take the pasilla dataset (in the DESeq2 vignette) as an example, if I am interested only in genes that show |log2 FC| >2, and I want to control for FDR <0.05, then I write:
dds <- DESeq(dds)
res <- results(dds, alpha = 0.05, lfcThreshold = 2, altHypothesis="greaterAbs")
resLFC <- lfcShrink(dds, coef = 2, res=res, lfcThreshold=2)
Is above code correct? In addition, do I need to put the lfcThreshold=2 argument in both results() step, and the lfcShrink() step?
I tried it on my computer, and the first few lines of the "resLFC" object (in the order of smallest FDR) look like below:
baseMean log2FoldChange lfcSE stat pvalue padj
FBgn0039155 730.567673 -3.686017982 0.12624706 -15.483906 4.455761e-54 4.420115e-50
FBgn0039827 261.911170 -2.841617881 0.14698260 -9.296996 1.444700e-20 7.165711e-17
FBgn0003360 4342.832062 -2.662356807 0.12047274 -8.215957 2.104796e-16 6.959859e-13
FBgn0025111 1501.447930 2.528934547 0.11059930 7.066198 1.592366e-12 3.893356e-09
FBgn0034736 225.870672 -2.492829764 0.14465009 -7.037131 1.962377e-12 3.893356e-09
FBgn0034434 114.623344 -2.227108812 0.15598919 -5.900835 3.616673e-09 5.310345e-06
FBgn0085359 68.606108 -1.856196452 0.16544745 -5.894982 3.747219e-09 5.310345e-06
FBgn0024288 58.849488 -1.911765311 0.16493679 -5.563957 2.637244e-08 3.270182e-05
How to interpret the last two genes(FBgn0085359, FBgn0024288) here? I mean, they both had significant FDR, but their |log2 FC| is not above my threshold of 2. So are they still significant DE genes?
I've studied the paper and the vignette, from them, I get the idea that log2 FC shrinkage is to overcome the low count-high variance issue, and is more preferable for ranking and visualization. So in a formal report(eg. publication), we should report the shrunken log2 FC, not the original log2 FC, is this correct?
As always, thank you so much for your time and explanation, Michael!
R version 3.4.3 (2017-11-30), DESeq2_1.18.1