DNA methylation analysis of ChIP products at single nucleotide resolution by Pyrosequencing®

Abstract

Interaction and co-occurrence of protein and DNA-based epigenetic modifications have become a topic of interest for many fundamental and biomedical questions. We describe within this chapter a protocol that combines two techniques in order to determine the methylation status of the DNA specifically associated with a protein of interest. First, DNA that directly interacts with the selected protein (such as a specific histone modification, a transcription factor, or any other DNA-associated protein) is purified by standard chromatin immunoprecipitation (ChIP). Second, the level of DNA methylation of this immunoprecipitated DNA is measured by bisulfite conversion and Pyrosequencing, a quantitative sequencing-by- synthesis method. This procedure allows determining the methylation status of genomic DNA associated to a specific protein at single nucleotide resolution.