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Updated: Sep 18, 2025
Single-Cell Factor Localization on Chromatin using Ultra-Low Input Cleavage Under Targets and Release using Nuclease
Published on: February 1, 2022
Cleavage Under Targets & Release Using Nuclease (CUT&RUN) Protocols for the Study of Genetic and Epigenetic
Stefan Custers1, Erika Apel2, Yujin Suk3
1Department of Biochemistry and Biomedical Sciences, Department of Surgery, McMaster Centre for Discovery in Cancer Research, McMaster University, Hamilton, ON, Canada.
Abstract:
Cancer cell malignancy is regulated by a variety of mechanisms, collectively termed the hallmarks of cancer. At the core of these regulative processes are genetic and epigenetic perturbations that play an essential role in neoplasmic formation. Advances in molecular techniques have enabled the in-depth research of the epigenome by means of epigenetic profiling, a process that has been dominated by the chromatin immunoprecipitation (ChIP) method. Even though ChIP has dominated the field, the technique is hindered by limitations such as high background noise, requirement of a large number of cells, requirement of antibodies with high pull-down efficiency, and crosslinking and solubilization-induced artifacts (e.g., epitope masking). Cleavage Under Targets and Release Under Nuclease (CUT&RUN) is a recently developed epigenomic alternative method to ChIP, offering advantages designed to overcome limitations of the ChIP method. Despite its advantages, it is not always straightforward which technique is most suitable for the job at hand. This chapter aims to provide an in-depth guide in performing CUT&RUN.
Insights
This guide details the CUT&RUN epigenomic profiling technique, an alternative to chromatin immunoprecipitation (ChIP). CUT&RUN offers advantages over ChIP for studying cancer epigenetics and neoplasm formation.
Area of Science:
- Molecular Biology
- Cancer Research
- Epigenetics
Background:
- Cancer cell malignancy is regulated by genetic and epigenetic mechanisms.
- Epigenetic profiling is crucial for understanding neoplasm formation.
- Chromatin immunoprecipitation (ChIP) has been the dominant method for epigenomic research.
Purpose of the Study:
- To provide an in-depth guide to performing the CUT&RUN technique.
- To highlight CUT&RUN as an alternative to ChIP for epigenomic profiling.
- To address the limitations of ChIP and the advantages of CUT&RUN.
Main Methods:
- The chapter focuses on the Cleavage Under Targets and Release Under Nuclease (CUT&RUN) method.
- CUT&RUN is presented as an alternative to Chromatin Immunoprecipitation (ChIP).
- The guide aims to facilitate the practical application of CUT&RUN.
Main Results:
- ChIP is limited by high background noise, large cell requirements, antibody dependency, and artifacts.
- CUT&RUN offers advantages over ChIP in epigenomic profiling.
- The chapter serves as a practical resource for researchers.
Conclusions:
- CUT&RUN is a valuable epigenomic profiling technique.
- Understanding the strengths of CUT&RUN compared to ChIP is essential for researchers.
- This guide aims to empower researchers to effectively utilize CUT&RUN.
