DMS‐seq for In Vivo Genome‐Wide Mapping of Protein‐DNA Interactions and Nucleosome Centers. Issue 1 (15th June 2018)

• Record Type: Journal Article
• Authors: Umeyama, Taichi; Ito, Takashi
• 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 123:Issue 1(2018)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2018-06-15
• Subjects: DNA‐binding protein -- epigenome -- genomic footprinting -- chromatin; 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/cpmb.60
• Languages: English
• ISSNs: 1934-3639

Abstract

Abstract: The genome exerts its functions through interactions with proteins. Hence, comprehensive identification of protein‐occupied sites by genomic footprinting is critical to an in‐depth understanding of genome functions. This unit describes the protocol of dimethyl sulfate‐sequencing (DMS‐seq). DMS is an alkylating reagent that methylates guanine and adenine in double‐stranded DNA. DMS added to the culture medium readily enters the cell and methylates its DNA throughout the genome except for the regions bound by proteins, thereby obviating the need for nuclear isolation in genomic footprinting. Polyamine/AP‐endonuclease treatment of DNA isolated from DMS‐treated cells induces cleavages at the methylated sites. Deep sequencing of these fragments identifies protein‐bound sites as peaks of protected fragments or troughs of cleavage sites. Furthermore, DMS displays an unexpected preference to nucleosome centers, enabling their direct detection without genetic manipulation. Therefore, DMS‐seq provides a unique method for non‐targeted profiling of in vivo protein‐DNA interactions. © 2018 by John Wiley & Sons, Inc.