TY - JOUR
T1 - Improved cut&run chromatin profiling tools
AU - Meers, Michael P.
AU - Bryson, Terri D.
AU - Henikoff, Jorja G.
AU - Henikoff, Steven
N1 - Funding Information:
We thank Christine Codomo and Tayler Hentges for technical support. We also thank all members of the Henikoff lab for valuable discussions and Kami Ahmad, Brian Freie and Bob Eisenman for comments on the manuscript. This work was supported by the Howard Hughes Medical Institute, and a grant from the National Institutes of Health (4DN TCPA A093) and the Chan-Zuckerberg Initiative.
Publisher Copyright:
© Meers et al.
PY - 2019
Y1 - 2019
N2 - 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.
AB - 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.
UR - http://www.scopus.com/inward/record.url?scp=85069236055&partnerID=8YFLogxK
U2 - 10.7554/ELIFE.46314
DO - 10.7554/ELIFE.46314
M3 - Article
C2 - 31232687
AN - SCOPUS:85071839202
SN - 2050-084X
VL - 8
JO - eLife
JF - eLife
M1 - e46314
ER -