Normalizing BAM Files

Question

[kstangline]

Hi!

I am back again with one more naive question.

What according to you is the best way to normalize multiple BAM files to one another?

My goal is to simply normalize a set of BAM files, then plot the avg read coverage.

I've attached an example of a read metagene plot showing the avg read coverage with BAM files that are not normalized to one another. To reiterate, my goal is to normalize these BAM files, then make the same type of plot.

[Ram]

Can you please define "normalize"? Do you mean to tweak the total number of aligned reads so all BAMs have comparable numbers of aligned reads?

[kstangline]

So the plot above is average read coverage, but I'm needing to make a new plot (the same type of figure as the one above) showing averaged normalized read coverage, which is normalized to the depth of the library.

Answer 1

[Devon Ryan]

You have a few options:

1. 1x normalize the BAM files with something like bamCoverage
2. Use multiBamSummary with the --scalingFactors to compute appropriate scaling factors in a more robust way and then use those to scale the per-BAM signals.