Simultaneous single-molecule detection of endogenous C-5 DNA methylation and chromatin accessibility using MAPit

Methods Mol Biol. 2012:833:125-41. doi: 10.1007/978-1-61779-477-3_9.

Abstract

Bisulfite genomic sequencing provides a single-molecule view of cytosine methylation states. After deamination, each cloned molecule contains a record of methylation within its sequence. The full power of this technique is harnessed by treating nuclei with an exogenous DNMT prior to DNA extraction. This exogenous methylation marks regions of accessibility and footprints nucleosomes, as well as other DNA-binding proteins. Thus, each cloned molecule records not only the endogenous methylation present (at CG sites, in mammals), but also the exogenous (GC, when using the Chlorella virus protein M.CviPI). We term this technique MAPit, methylation accessibility protocol for individual templates.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • 5-Methylcytosine / metabolism*
  • Base Sequence
  • Chromatin / metabolism*
  • Chromatin Assembly and Disassembly
  • Cloning, Molecular
  • DNA Methylation / genetics*
  • Humans
  • K562 Cells
  • Molecular Biology / methods*
  • Molecular Sequence Data
  • Sulfites
  • Transcription Initiation Site

Substances

  • Chromatin
  • Sulfites
  • 5-Methylcytosine
  • hydrogen sulfite