Improved CUT&RUN chromatin profiling tools

Elife. 2019 Jun 24;8:e46314. doi: 10.7554/eLife.46314.

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.

Keywords: chromatin; chromosomes; epigenomics; gene expression; genetics; genomics; human; spike-in calibration.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Chromatin / metabolism*
  • Epigenesis, Genetic
  • Immunologic Techniques / methods*
  • Micrococcal Nuclease / genetics
  • Micrococcal Nuclease / metabolism
  • Molecular Biology / methods*
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Staphylococcal Protein A / genetics
  • Staphylococcal Protein A / metabolism

Substances

  • Bacterial Proteins
  • Chromatin
  • IgG Fc-binding protein, Streptococcus
  • Recombinant Fusion Proteins
  • Staphylococcal Protein A
  • Micrococcal Nuclease

Associated data

  • GEO/GSE126612