RADICL-ChIP: a Method to Capture Global RNA-on-Genome Distribution Mediated by Chromatin associated Factors

Abstract

Chromatin associated RNAs play a key role in mediating epigenetic mechanism. To improve our comprehension of how chromatin-associated RNAs influence gene expression, it is crucial to identify the genomic locus that RNA interacts with on a genome-wide scale. Emerging technologies were developed to capture binding sites of lncRNAs globally. Such techniques rely on the concept of Proximity ligation via a biotinylated adapter to ligate DNA and RNA on each end, such as MARGI, GRID-seq, ChAR-seq and the most recent technology, devised in our lab RNA And DNA Complexes Ligated & Sequenced (RADICL-seq). RADICL-seq has several advantages over the other methods. However, while this method produced a fairly global picture of the chromatin associated “RNA-ome”, the specific association with specific regulatory factors is missing, in addition all emerging technologies lack the sensitivity to capture low-expressed RNAs. Thus, we set out a strategy to address this issue by enriching global RNA-chromatin interactions mediated by a specific factor which will be achieved by combining RADICL with Chromatin Immunoprecipitation (ChIP) method. Since our lab interest is investigating the role of lncRNAs in muscle differentiation, RADICL-ChIP has been performed using the C2C12 mouse skeletal muscle. In particular, the role of identification of RNA moieties interacting with PRC2 PcG proteins in stress response and their genome wide distribution in chromatin.