Normalizing Bedgraphs for comparisons between sequencing experiments?

Question

[albert.dayuan.yu]

Hello all,

I'm trying to compare coverage within specific introns across different RNA-seq experiments, and I was hoping to do so through bedgraph files. Would deeptools bamCoverage normalization be adequate for enabling comparisons between bedgraphs? I've been doing so using the normalizeTo1x flag.

Answer 1

[ATpoint]

You could use the normalization factors produced by e.g. edgeR to scale the raw counts. For this you would need to calculate norm. factors in R using a standard count matrix e.g. with the exon counts (you could use featureCounts for creation of the matrix directly from the BAM files) and then use edgeR for normalization followed by providing these factors to bamCoverage. See the link below for example code:

A: ATAC-seq sample normalization (quantil normalization)