Improved CUT&RUN chromatin profiling tools
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 enha...
| Main Authors: | , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
eLife Sciences Publications Ltd
2019-06-01
|
| Series: | eLife |
| Subjects: | |
| Online Access: | https://elifesciences.org/articles/46314 |
| _version_ | 1818019938661564416 |
|---|---|
| author | Michael P Meers Terri D Bryson Jorja G Henikoff Steven Henikoff |
| author_facet | Michael P Meers Terri D Bryson Jorja G Henikoff Steven Henikoff |
| author_sort | Michael P Meers |
| collection | DOAJ |
| description | 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. |
| first_indexed | 2024-04-14T07:58:23Z |
| format | Article |
| id | doaj.art-75191d395f574c6ea9d1e96c5b479ffb |
| institution | Directory Open Access Journal |
| issn | 2050-084X |
| language | English |
| last_indexed | 2024-04-14T07:58:23Z |
| publishDate | 2019-06-01 |
| publisher | eLife Sciences Publications Ltd |
| record_format | Article |
| series | eLife |
| spelling | doaj.art-75191d395f574c6ea9d1e96c5b479ffb2022-12-22T02:04:58ZengeLife Sciences Publications LtdeLife2050-084X2019-06-01810.7554/eLife.46314Improved CUT&RUN chromatin profiling toolsMichael P Meers0Terri D Bryson1Jorja G Henikoff2Steven Henikoff3https://orcid.org/0000-0002-7621-8685Basic Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, United StatesBasic Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, United States; Howard Hughes Medical Institute, United StatesBasic Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, United StatesBasic Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, United States; Howard Hughes Medical Institute, United StatesPreviously, 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.https://elifesciences.org/articles/46314chromatinepigenomicsspike-in calibration |
| spellingShingle | Michael P Meers Terri D Bryson Jorja G Henikoff Steven Henikoff Improved CUT&RUN chromatin profiling tools eLife chromatin epigenomics spike-in calibration |
| title | Improved CUT&RUN chromatin profiling tools |
| title_full | Improved CUT&RUN chromatin profiling tools |
| title_fullStr | Improved CUT&RUN chromatin profiling tools |
| title_full_unstemmed | Improved CUT&RUN chromatin profiling tools |
| title_short | Improved CUT&RUN chromatin profiling tools |
| title_sort | improved cut run chromatin profiling tools |
| topic | chromatin epigenomics spike-in calibration |
| url | https://elifesciences.org/articles/46314 |
| work_keys_str_mv | AT michaelpmeers improvedcutrunchromatinprofilingtools AT terridbryson improvedcutrunchromatinprofilingtools AT jorjaghenikoff improvedcutrunchromatinprofilingtools AT stevenhenikoff improvedcutrunchromatinprofilingtools |