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Multidimensional chemical control of CRISPR–Cas9

Abstract

Cas9-based technologies have transformed genome engineering and the interrogation of genomic functions, but methods to control such technologies across numerous dimensions—including dose, time, specificity, and mutually exclusive modulation of multiple genes—are still lacking. We conferred such multidimensional controls to diverse Cas9 systems by leveraging small-molecule-regulated protein degron domains. Application of our strategy to both Cas9-mediated genome editing and transcriptional activities opens new avenues for systematic genome interrogation.

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Figure 1: Multidimensional 'chemical' control of endogenous transcript levels.
Figure 2: Dose and temporal regulation of DD.SpCas9.DD-mediated insertion/deletion (indel) formation.

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Acknowledgements

This work was supported by the NIH (Director's New Innovator Award 1DP2GM119162 to M.D.S.; grant 1R21AI126239-01 to A.C.), the Edward Mallinckrodt, Jr. Foundation (Faculty Scholar Award to M.D.S.), and the Burroughs Wellcome Fund (Career Award at the Scientific Interface to A.C.). C.L.M. acknowledges the National Science Foundation for a Graduate Research Fellowship. We are grateful to B. Harvey (NIDA) and to I. Slaymaker, F.A. Ran, and B. Wagner (Broad Institute) for helpful discussions. J.K. Joung (Harvard Medical School, Boston, Massachusetts, USA) provided U2OS.eGFP-PEST cells. This work is dedicated to Professor Stuart L. Schreiber on the occasion of his 60th birthday.

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B.M., C.L.M., B.Z., M.D.S., and A.C. planned research and analyzed data; B.M., C.L.M., B.Z., F.Z., M.D.S., and A.C. designed experiments; B.M., C.L.M., B.Z., and S.E.V. performed experiments; B.M., C.L.M., M.D.S., and A.C. wrote the manuscript; M.D.S. and A.C. supervised research.

Corresponding authors

Correspondence to Matthew D Shoulders or Amit Choudhary.

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Broad Institute has filed a patent application including work from this paper.

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Supplementary Results, Supplementary Tables 1–5, Supplementary Figures 1–13 and Supplementary Note. (PDF 3762 kb)

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Maji, B., Moore, C., Zetsche, B. et al. Multidimensional chemical control of CRISPR–Cas9. Nat Chem Biol 13, 9–11 (2017). https://doi.org/10.1038/nchembio.2224

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