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Elife. 2017 Jan 16;6. pii: e21856. doi: 10.7554/eLife.21856.

An efficient targeted nuclease strategy for high-resolution mapping of DNA binding sites.

Author information

1
Howard Hughes Medical Institute, Basic Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, United States.

Abstract

We describe Cleavage Under Targets and Release Using Nuclease (CUT&RUN), a chromatin profiling strategy in which antibody-targeted controlled cleavage by micrococcal nuclease releases specific protein-DNA complexes into the supernatant for paired-end DNA sequencing. Unlike Chromatin Immunoprecipitation (ChIP), which fragments and solubilizes total chromatin, CUT&RUN is performed in situ, allowing for both quantitative high-resolution chromatin mapping and probing of the local chromatin environment. When applied to yeast and human nuclei, CUT&RUN yielded precise transcription factor profiles while avoiding crosslinking and solubilization issues. CUT&RUN is simple to perform and is inherently robust, with extremely low backgrounds requiring only ~1/10th the sequencing depth as ChIP, making CUT&RUN especially cost-effective for transcription factor and chromatin profiling. When used in conjunction with native ChIP-seq and applied to human CTCF, CUT&RUN mapped directional long range contact sites at high resolution. We conclude that in situ mapping of protein-DNA interactions by CUT&RUN is an attractive alternative to ChIP-seq.

KEYWORDS:

DNA sequencing; S. cerevisiae; chromatin; chromosomes; genes; human; in situ profiling; transcription factors

Comment in

PMID:
28079019
PMCID:
PMC5310842
DOI:
10.7554/eLife.21856
[Indexed for MEDLINE]
Free PMC Article

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