Question: Possible reason for no peaks generated by MACS2
gravatar for Louis Kok
14 months ago by
Louis Kok10
Louis Kok10 wrote:

Hi, I am new to ChIP-Seq data analysis. In my samples, the expression of transcription factor of interest was observed to be down-regulated. Subsequently, a ChIP-Seq on this transcription factor of interest in three replicates was performed and input DNA was used as control. Initially when I performed MACS2 analysis using the command below:

mac2 callpeak -t treatment.rep1.bam treatment.rep2.bam treatmen.rep3.bam -c input.rep1.bam input.rep2.bam input.rep3.bam -f BAM -g hs -n output.macs2

The run failed and I received the error below:

WARNING @ Wed, 10 Oct 2018 11:53:34: Too few paired peaks (0) so I can not build the model! Broader your MFOLD range parameter may erase this error. If it still can't build the model, we suggest to use --nomodel and --extsize 147 or other fixed number instead.

After adding --nomodel to the command. The run was successful but not peak was outputted.

My question is: Is it expected for no peak to be detected for samples with down-regulated TF? Or should I further optimize the parameters to increase the chance to get some peaks?

I was wondering if there might be at least some peaks detected due to noise.

Thanks in advance.

chip-seq macs • 947 views
ADD COMMENTlink modified 13 months ago by Biostar ♦♦ 20 • written 14 months ago by Louis Kok10

If you heavily knock-down TF expression it's unsurprising that there are no peaks. I wonder, how well the library prep worked. Were you able to get decent amounts of DNA from the IP still? Have you looked at a fingerprint from plotFingerprint? If you got a decent amount of DNA still and the fingerprint looks OK then often playing around with --slocal and --llocal helps.

ADD REPLYlink written 14 months ago by Devon Ryan93k


You should maybe start with generating coverage files (wig, tdf, bedgraph...) and checking what your samples look like with a genome browser, see if it looks alright.

As for Macs2, as it says in the error message: Broader your MFOLD range parameter may erase this error. By default, the mfold range is 5-50, maybe you can lower the threshold and see what happens:

   macs2 callpeak --mfold 2 50 ...
ADD REPLYlink modified 13 months ago • written 13 months ago by rioualen390

Can you be sure that the antibody is of high quality and able to do decent IP if true peaks were present.

ADD REPLYlink written 13 months ago by ATpoint26k
Please log in to add an answer.


Use of this site constitutes acceptance of our User Agreement and Privacy Policy.
Powered by Biostar version 2.3.0
Traffic: 907 users visited in the last hour