Improved CUT&RUN chromatin profiling tools
- Fred Hutchinson Cancer Research Center, United States
Abstract
Previously we described a novel alternative to Chromatin Immunoprecipitation, CUT&RUN, in which unfixed permeabilized cells are incubated with antibody, followed by binding of a Protein A-Micrococcal Nuclease (pA/MNase) fusion protein (Skene and Henikoff, 2017). Here we introduce three enhancements to CUT&RUN: A hybrid Protein A-Protein G-MNase construct that expands antibody compatibility and simplifies purification, a modified digestion protocol that inhibits premature release of the nuclease-bound complex, and a calibration strategy based on carry-over of E. coli DNA introduced with the fusion protein. These new features, coupled with the previously described low-cost, high efficiency, high reproducibility and high-throughput capability of CUT&RUN make it the method of choice for routine epigenomic profiling.
Data availability
The plasmid pAG-ERH-MNase-6xHIS-HA is available from Addgene. Sequencing datasets are available from GEO (GSE126612).
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A streamlined protocol and analysis pipeline for CUT&RUN chromatin profilingNCBI Gene Expression Omnibus, GSE126612.
Article and author information
Author details
Funding
Howard Hughes Medical Institute
- Michael P Meers
National Institutes of Health (4DN TCPA A093)
- Terri D Bryson
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2019, Meers et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Improved CUT&RUN chromatin profiling tools
eLife 8:e46314.
https://doi.org/10.7554/eLife.46314
