journal.pone.0133246.PDF (2.93 MB)
Retention of the native epigenome in purified mammalian chromatin
journal contributionposted on 2020-07-21, 10:41 authored by Andreas H Ehrensberger, Don-Marc Franchini, Philip East, Roger George, Nik Matthews, Sarah L Maslen, Jesper Q Svejstrup
A protocol is presented for the isolation of native mammalian chromatin as fibers of 25-250 nucleosomes under conditions that preserve the natural epigenetic signature. The material is composed almost exclusively of histones and DNA and conforms to the structure expected by electron microscopy. All sequences probed for were retained, indicating that the material is representative of the majority of the genome. DNA methylation marks and histone marks resembled the patterns observed in vivo. Importantly, nucleosome positions also remained largely unchanged, except on CpG islands, where nucleosomes were found to be unstable. The technical challenges of reconstituting biochemical reactions with native mammalian chromatin are discussed.