RNA-programmed genome editing in human cells

Abstract

Type II CRISPR immune systems in bacteria use a dual RNA-guided DNA endonuclease, Cas9, to cleave foreign DNA at specific sites. We show here that Cas9 assembles with hybrid guide RNAs in human cells and can induce the formation of double-strand DNA breaks (DSBs) at a site complementary to the guide RNA sequence in genomic DNA. This cleavage activity requires both Cas9 and the complementary binding of the guide RNA. Experiments using extracts from transfected cells show that RNA expression and/or assembly into Cas9 is the limiting factor for Cas9-mediated DNA cleavage. In addition, we find that extension of the RNA sequence at the 3′ end enhances DNA targeting activity in vivo. These results show that RNA-programmed genome editing is a facile strategy for introducing site-specific genetic changes in human cells.

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• The molecular basis for recognition of 5′-NNNCC-3′ PAM and its methylation state by Acidothermus cellulolyticus Cas9 · 2020
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• Ex Vivo/In vivo Gene Editing in Hepatocytes Using “All-in-One” CRISPR-Adeno-Associated Virus Vectors with a Self-Linearizing Repair Template · 2020
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• The promise and challenge of therapeutic genome editing · 2020
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• Massively parallel profiling and predictive modeling of the outcomes of CRISPR/Cas9-mediated double-strand break repair · 2019
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• A Stem-Cell-Derived Platform Enables Complete Cryptosporidium Development In Vitro and Genetic Tractability · 2019
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• Stammzellforschung · 2018
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• The Current State and Future of CRISPR-Cas9 gRNA Design Tools · 2018
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• Transgenic Mouse Models in Cancer Research · 2018
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and 42 more

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