For Chip-seq, which control data I should choose for each biological replicate samples in MACS2

Question

[bxia]

I have control and experimental groups, each has 3 biological replicates.

When I do the Call peaks in MACS2, how to I pick the combination?

if I combine all the experimental files and control files, the output will be combined all of them and I will lose my biological replicates?

Should I just pick one to analyze?

Thanks

[SP]

I have to ask, do you want to check if the peaks are lost or gained wrt control in exprimental group?

Answer 1

[ivivek_ngs]

1. You can take a look at IDR that was used for ENCODE.

2. Take a look at this link for the tools and the approaches highlighted and also here. I would suggest latest version of MACS2 if you are using it for farther analysis.

3. You can also use deeptools to find the correlation of your bam files both genome wide or promoter wide to see the discrepancy and judge if you want to proceed with differential binding or not.

4. I would personally suggest for running in parallel the peak calling for control/ input experimenta/input as am guessing you have such an experimental designl for each samples and then perform a multipeak overlap with either bedtools or HOMER and then annotate them to find the regions of interest and proceed for downstream exploratory analysis. You can also do the task of differential binding with diffreps/Pepr/MACS2 to assess regions that have differential binding between control and experimental conditions and then highlight one the regions with IGV or any standard genomic browser.

Good luck!

Answer 2

[James Ashmore]

Usually the test and control samples are prepared in tandem, so I would suggest you use replicate 1 of the test with replicate 1 of the control e.t.c