Targeted in situ genome-wide profiling with high efficiency for low cell numbers
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Abstract
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.
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Recent methods have overcome some limitations of ChIP.
“While ChIP is not equally adept at measuring all interactions (Baranello et al., 2016; Teytelman et al., 2013), recent methods have overcome some of these limitations (Janssens et al., 2018; Skene and Henikoff, 2017; Skene et al., 2018; Zhu et al., 2019).”
From Unraveling Hematopoiesis through the Lens of Genomics (2020) · cites this paper as Skene et al., 2018
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- Authors
- Skene, P. J., Henikoff, J. G., & Henikoff, S.
- Affiliations
- Howard Hughes Medical Institute (US)
- Fred Hutch Cancer Center (US)
- Corresponding author
- Henikoff, Steven
- Venue
- Nature Protocols
- Published
- 2018-04-12
- Volume
- 13
- Issue
- 5
- Pages
- 1006-1019
- ISSN
- 1750-2799 (ISSN-L)
- 1754-2189
- OpenAlex
- W2757704052
- Cited by
- 803
- References
- 50
- Citations by year
- 2026: 69 · 2025: 112 · 2024: 130 · 2023: 97 · 2022: 127 · 2021: 119 · 2020: 78 · 2019: 54 · 2018: 15 · 2017: 1 · 2016: 1
- FWCI
- 23.65
- Type
- journal article
- Language
- en
- Open access
- green
- doi.org
- Licence
- cc-by-nc
- www.springer.com/tdm (tdm)
- Locations
- Nature Protocols (publishedVersion)
- bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Chromatin Dynamics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Chromatin Remodeling and Cancer
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Epigenetics and DNA Methylation
- Keywords
- CUT&RUN, staphylococcal nuclease, protein-DNA complex, histone modifications, transcription factors, epigenetic profiling, signal-to-noise ratio, protocol optimization
- MeSH terms
- Animals, Cell Line, Chromatin, DNA, DNA-Binding Proteins, Humans, Epigenomics
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