Targeted in situ genome-wide profiling with high efficiency for low cell numbers

By P. Skene, Jorja G. Henikoff, Steven Henikoff. Nature Protocols, 2018

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This protocol describes CUT&RUN, a genome-wide targeted epigenetic profiling method that allows the analysis of low cell numbers at high resolution. Cleavage under targets and release using nuclease (CUT&RUN) is an epigenomic profiling strategy in which antibody-targeted controlled cleavage by micrococcal nuclease releases specific protein–DNA complexes into the supernatant for paired-end DNA sequencing. As only the targeted fragments enter into solution, and the vast majority of DNA is left behind, CUT&RUN has exceptionally low background levels. CUT&RUN outperforms the most widely used chromatin immunoprecipitation (ChIP) protocols in resolution, signal-to-noise ratio and depth of sequencing required. In contrast to ChIP, CUT&RUN is free of solubility and DNA accessibility artifacts and has been used to profile insoluble chromatin and to detect long-range 3D contacts without cross-linking. Here, we present an improved CUT&RUN protocol that does not require isolation of nuclei and provides high-quality data when starting with only 100 cells for a histone modification and 1,000 cells for a transcription factor. From cells to purified DNA, CUT&RUN requires less than a day at the laboratory bench and requires no specialized skills.

Subfields: Molecular Biology

View Paper: https://www.semanticscholar.org/paper/cf64a876d302225e8b72ff8bf2dfb77214ac4227

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