Question: Problems with MACS ChIP-Seq peak calling
0
gravatar for dcheng1
22 months ago by
dcheng10
dcheng10 wrote:

I downloaded raw data in GEO and analyzed it following general ChIP-Seq pipeline: trimming adapter, mapping and calling peaks. However, when compared with bigwig files in GEO, my results look like separate spikes instead of peaks.

Here is the command I used: macs14 --bw 200 -t -c -B -S --call-subpeaks

Does anyone know how did this happen? Thanks!!

enter image description here

chip-seq peak calling macs • 951 views
ADD COMMENTlink modified 22 months ago by YaGalbi1.4k • written 22 months ago by dcheng10
1

Need more information

We need to know the MACS2 output file you are working from. Do you load this file directly into IGV or create another file from loading it? Always include file extensions Please print the top 10 lines (including header if there is one) of the file you are loading into IGV

ADD REPLYlink modified 22 months ago • written 22 months ago by YaGalbi1.4k
track type=bedGraph name="NA_treat_all" description="Extended tag pileup from MACS version 1.4.2 20120305"
chr1    10002   10003   1
chr1    10003   10004   2
chr1    10004   10008   3
chr1    10008   10014   4
chr1    10014   10015   5
chr1    10015   10016   7
chr1    10016   10020   8
chr1    10020   10021   9
chr1    10021   10026   10
ADD REPLYlink modified 22 months ago by WouterDeCoster38k • written 22 months ago by dcheng10
1

you need to create a signal track. Calling peaks is one thing, visualising them is something else. MACS2 will have created 2 .bdg file called [prefix]_treat_pileup.bdg and [prefix]_control_lambda.bdg for your sample. These are used to create the type of file you want. The link describes how to create a FE or LogLR track but I think you need a ppois track.

$ macs2 bdgcmp -t [prefix]_treat_pileup.bdg -c [prefix]_control_lambda.bdg -o [prefix]_ppois.bdg -m ppois
ADD REPLYlink modified 22 months ago • written 22 months ago by YaGalbi1.4k
1

I added markup to your post for increased readability. You can do this by selecting the text and clicking the 101010 button. When you compose or edit a post that button is in your toolbar, see image below:

101010 Button

ADD REPLYlink written 22 months ago by WouterDeCoster38k

Link for igv screenshot: https://ibb.co/dXZhvv

ADD REPLYlink written 22 months ago by dcheng10

MACS call peaks (regions where there are peaks). BigWigs are just the coverage tracks (at least in this case). Those should not look similar.

ADD REPLYlink written 22 months ago by igor7.6k
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