I have recently performed some ChIP experiments and did qPCR for GAPDH promoter to check the specificity of the ChIP in RNA pol compared to IgG sample.
qPCR showed 2 fold enrichment of GAPDH promoter in RNA pol sample compared to IgG sample.
My specific question is; if 2 fold enrichment is acceptable or not? what is the threshold of that to consider a ChIP experiment specific enough to go for further steps of library prep and sequencing?
The specific TF which I am trying to do ChIP on, has no known target genes. So, there is no option for me to do qPCR on positive or negative binding regions of that gene to check the specificity.
I also did qPCR for GAPDH on the TF ChIPed DNA and the Ct value was near what I got for IgG sample. All these show that probably there is a specificity but the difference is quite low (only 2 fold). ( See the attached photo)
Can anyone help me in what is considered as the threshold for fold enrichment?