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2 hours ago by

I am analyzing chip-seq data for several DNA binding proteins and doing differential binding analysis between two different conditions. I am using HOMER for differential analysis which basically uses the standard Deseq2 pipeline to compare counts between peak regions. Using mac2 gives me ~20,000 peaks for each protein of interest but when I do differential analysis, none of these peaks show differential binding with FDR cutoff 5% between my treatment conditions. This is very unexpected because the proteins are expected to lose binding in my treatment condition. I am suspicious that I don’t even get a single differential peak out of ~20,000 peaks in three different chip-seq datasets.

Looking at the p-values of my differential analysis shows me the there are hundreds of peaks with significant unadjusted p-values but all of them have very high adjusted p-values. Also, almost all peaks have identical adjusted p-value of ~0.98 which is very strange. After reading previous threads about this I made a histogram of my unadjusted p-vales which is not uniform but upward sloping (similar to scenario D in this article).

I am at a loss for what could be causing this and would appreciate any tips.

Thanks



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