QuestionsCategory: ChIP-Seq Data AnalysisControl Types: Why does my IgG control sample have higher binding than the antibody binding sample for qPCR? What do I do about it?
stonelakeak@nih.gov Staff asked 5 months ago

Why does my IgG control sample have higher binding than the antibody binding sample for qPCR? What do I do about it?

1 Answers
stonelakeak@nih.gov Staff answered 5 months ago

There will always be sites in the genome where the control sample has higher binding than the ChIP. For an IgG sample, this is caused by the antibody having fewer places in the genome to bind relative to the ChIP causing an increase in its binding at non-specific sites and a higher binding in the control than the ChIP. If this is a site you are trying to use for qPCR, it may not be the optimal location. If you can, you should probably look for an alternative. For the purposes of ChIP-seq, just don’t focus on that region of the genome for your analysis if at all possible and make sure you are using your control in your analyses. – answered by Tovah Markowitz, Paul Schaughency, Vishal Koparde