RNA antisense purification (RAP) for mapping RNA interactions with chromatin

Methods Mol Biol. 2015:1262:183-97. doi: 10.1007/978-1-4939-2253-6_11.

Abstract

RNA-centric biochemical purification is a general approach for studying the functions and mechanisms of noncoding RNAs. Here, we describe the experimental procedures for RNA antisense purification (RAP), a method for selective purification of endogenous RNA complexes from cell extracts that enables mapping of RNA interactions with chromatin. In RAP, the user cross-links cells to fix endogenous RNA complexes and purifies these complexes through hybrid capture with biotinylated antisense oligos. DNA loci that interact with the target RNA are identified using high-throughput DNA sequencing.

MeSH terms

  • Animals
  • Cells, Cultured
  • Chromatin / metabolism*
  • DNA / metabolism*
  • Gene Library
  • High-Throughput Nucleotide Sequencing
  • Mice
  • RNA, Antisense / isolation & purification*
  • Sequence Analysis, RNA
  • Software

Substances

  • Chromatin
  • RNA, Antisense
  • DNA