Cas9 gRNA engineering for genome editing, activation and repression

Nat Methods. 2015 Nov;12(11):1051-4. doi: 10.1038/nmeth.3580. Epub 2015 Sep 7.


We demonstrate that by altering the length of Cas9-associated guide RNA (gRNA) we were able to control Cas9 nuclease activity and simultaneously perform genome editing and transcriptional regulation with a single Cas9 protein. We exploited these principles to engineer mammalian synthetic circuits with combined transcriptional regulation and kill functions governed by a single multifunctional Cas9 protein.

Publication types

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

MeSH terms

  • Binding Sites
  • CRISPR-Associated Proteins / genetics
  • CRISPR-Cas Systems / genetics*
  • Flow Cytometry
  • Fluorescent Dyes / analysis
  • Gene Deletion
  • Genes, Reporter
  • Genetic Engineering / methods
  • Genetic Vectors
  • Genome
  • HEK293 Cells
  • Humans
  • Microscopy, Fluorescence
  • Mutagenesis
  • Mutation
  • RNA Editing
  • RNA, Guide, CRISPR-Cas Systems
  • Transcription, Genetic


  • CRISPR-Associated Proteins
  • Fluorescent Dyes