MARCC (Matrix‐Assisted Reader Chromatin Capture): An Antibody‐Free Method to Enrich and Analyze Combinatorial Nucleosome Modifications. Issue 1 (1st July 2015)

• Record Type: Journal Article
• Authors: Su, Zhangli; Denu, John M.
• Editors: Ausubel, Frederick M.; Brent, Roger; Kingston, Robert E.; Moore, David D.; Seidman, J.G.; Smith, John A.; Struhl, Kevin
• Is Part Of: Current protocols in molecular biology. Volume 111:Issue 1(2015)
• Page Start: 21.32.1
• Page End: 21.32.21
• Publication Date: 2015-07-01
• Subjects: affinity enrichment -- antibody‐free -- chromatin -- histone -- PTM -- reader domain -- MARCC; Molecular biology -- Technique -- Periodicals; Molecular biology -- Laboratory manuals; Molecular Biology -- methods; Biologie moléculaire -- Technique; Biologie moléculaire -- Manuels de laboratoire; Molecular biology; Molecular biology -- Technique; Laboratory Manual; Electronic reference sources; Laboratory manuals; 572.8028
• DOI: 10.1002/0471142727.mb2132s111
• Languages: English
• ISSNs: 1934-3639

Abstract

Abstract: Combinatorial patterns of histone modifications are key indicators of different chromatin states. Most of the current approaches rely on the usage of antibodies to analyze combinatorial histone modifications. Here we detail an antibody‐free method named MARCC (M atrix‐A ssisted R eader C hromatin C apture) to enrich combinatorial histone modifications. The combinatorial patterns are enriched on native nucleosomes extracted from cultured mammalian cells and prepared by micrococcal nuclease digestion. Such enrichment is achieved by recombinant chromatin‐interacting protein modules, or so‐called reader domains, which can bind in a combinatorial modification‐dependent manner. The enriched chromatin can be quantified by immunoblotting or mass spectrometry for the co‐existence of histone modifications, while the associated DNA content can be analyzed by qPCR or next‐generation sequencing. Altogether, MARCC provides a reproducible, efficient and customizable solution to enrich and analyze combinatorial histone modifications. © 2015 by John Wiley & Sons, Inc.