Journal
ELIFE
Volume 2, Issue -, Pages -Publisher
ELIFE SCIENCES PUBLICATIONS LTD
DOI: 10.7554/eLife.00471
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Funding
- Howard Hughes Medical Institute
- National Institutes of Health [R01 GM65462]
- Direct For Biological Sciences [950971] Funding Source: National Science Foundation
- Div Of Molecular and Cellular Bioscience [950971] Funding Source: National Science Foundation
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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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