Work

Mapping Regulatory Factors by Immunoprecipitation from Native Chromatin

Year: 2015

Type: article

Abstract: Occupied Regions of Genomes from Affinity-purified Naturally Isolated Chromatin (ORGANIC) is a high-resolution method that can be used to quantitatively map protein-DNA interactions with high specificity and sensitivity. This method uses micrococcal nuclease (MNase) digestion of chromatin and low-salt solubilization to preserve protein-DNA complexes, followed by immunoprecipitation and paired-end sequencing for genome-wide mapping of binding sites. In this unit, we describe methods for isolation of nuclei and MNase digestion of unfixed chromatin, immunoprecipitation of protein-DNA complexes, and high-throughput sequencing to map sites of bound factors.

Source: Current Protocols in Molecular Biology

DOI: 10.1002/0471142727.mb2131s110

Authors: Guillermo A. Orsi, Sivakanthan Kasinathan, Gabriel E. Zentner, Steven Henikoff, Kami Ahmad