Question: Downsampling ChIP-seq BAM files with spike in normalization factor before feeding into Diffbind
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gravatar for sikhtechai
3 months ago by
sikhtechai0
sikhtechai0 wrote:

I am planning to use exogenous chromatin as a spike in control with my actual sample from mouse to perform ChIP-seq for peak calling and differential binding analysis for histone modifications. this involves down-sampling the uniquely mapped read files to the calculated normalization factor from the spike in. This is seemingly helpful for peak calling and visualizing in genome browser. But I have not found any reference on whether it is considered in differential binding analysis. Since I am using Diffbind, I also could not find anything regarding this in the vignette. Could anyone please explain how this strategy might affect using the diffbind package? Or, does using spike-in for ChIP-seq normalization makes sense?

I greatly appreciate your time to read and answer to my question! Thank you in advance!

An example of this normalization process is given below(Image from Active motif's ChIP-seq spike in kit.) From active motif

ADD COMMENTlink modified 3 months ago by Jennifer Hillman Jackson25k • written 3 months ago by sikhtechai0
0
gravatar for Jennifer Hillman Jackson
3 months ago by
United States
Jennifer Hillman Jackson25k wrote:

Hello,

I would suggest using the Bioconductor/Diffbind support forum for questions about novel protocols if you cannot find literature references.

For example, this is prior Q&A based off a few keywords in your question: https://support.bioconductor.org/local/search/page/?q=Diffbind+normalization

Scientific usage in Galaxy is the same as when used line command for this and most 3rd party wrapped tools. Parameter options are labeled with the command-line parameter flag on the tool form.

Galaxy tutorials: https://galaxyproject.org/learn/

Thanks! Jen, Galaxy team

ADD COMMENTlink modified 3 months ago • written 3 months ago by Jennifer Hillman Jackson25k
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