Isolation of DNA by EdU-labelling for Mass Spectrometry (iDEMS)

Abstract DNA methylation and hydroxymethylation are important regulators of gene expression and genome stability in mammalian genomes. DNA replication dilutes these marks, and their restoration kinetics are important to epigenetic cell memory. We therefore developed isolation of DNA by EdU-labelling for Mass Spectrometry (iDEMS), a highly sensitive, quantitative mass spectrometry-based method for measuring DNA modifications on metabolically labelled DNA. Unlike sequencing-based methods, iDEMS does not require any amplification step, providing direct measurements from labelled DNA. Through additional methionine-isotope labelling and/ or incorporation of downstream steps such as chromatin immunoprecipitation, iDEMS facilitates novel insights into methylation metabolism and epigenetic crosstalk in chromatin. Excluding cell culture, DNA labelling and isolation requires two working days including an overnight digestion; preparation and running of samples requires three to four days. This protocol is related to the publication “Quantifying propagation of DNA methylation and hydroxymethylation with iDEMS” in Nature Cell Biology.