Calculating nucleosome dyad densities

Question

[mmmmcandrew]

Hi all- I would like to calculate nucleosome dyad densities from my MNase-seq data. Here is where things get complicated: I would first like to take the ratio of two normalized datasets, then calculate the dyad density from there. I have been successful in producing ratio files from two normalized datasets using deepTools bamCompare, but this can only generate BigWig or BedGraph files. I would now like to calculate dyad densities from those files. Is there a way this can be done? Are there alternative programs which would allow me to take the ratio of normalized datasets and produce a bam file as a ratio instead? Producing a BAM file would allow me to use deepTools bamCoverage "--MNase" option. Forgive me if this seems a stupid question, the concept of dyad density calculation is pretty foreign to me.

[Devon Ryan]

You can use bamCoverage with the --MNase function first and then bigwigCompare the results to presumably end up with your desired bigWig track.

[mmmmcandrew]

Cool, thanks for the input. This is kind of where I was thinking of going. My only concern is that I would prefer to normalize the datasets and take the ratio first, but maybe this can't be helped.

[Devon Ryan]

There's no way to go from bigWig->BAM, making a bigWig file is innately a lossy process. I would recommend that you do the following:

1. Run bamCompare, but note the scale factors applied to each sample. You might need to use the --verbose option, I don't recall.
2. Run bamCoverage --MNase on each sample to make bigWig files.
3. Run bigwigCompare with the --scaleFactors set according to step 1. This should produce more or less exactly what you're trying to get (at least I suspect so).