Targeted genome engineering in human cells with the Cas9 RNA-guided endonuclease

We employ the CRISPR-Cas system of Streptococcus pyogenes as programmable RNA-guided endonucleases (RGENs) to cleave DNA in a targeted manner for genome editing in human cells. We show that complexes of the Cas9 protein and artificial chimeric RNAs efficiently cleave two genomic sites and induce indels with frequencies of up to 33%.

[1]  Eunji Kim,et al.  Targeted chromosomal deletions in human cells using zinc finger nucleases. , 2010, Genome research.

[2]  Seung Woo Cho,et al.  Targeted genome editing in human cells with zinc finger nucleases constructed via modular assembly. , 2009, Genome research.

[3]  J. Doudna,et al.  A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity , 2012, Science.

[4]  Le Cong,et al.  Multiplex Genome Engineering Using CRISPR/Cas Systems , 2013, Science.

[5]  J. Keith Joung,et al.  FLASH Assembly of TALENs Enables High-Throughput Genome Editing , 2012, Nature Biotechnology.

[6]  Jin-Soo Kim,et al.  Genome editing with modularly assembled zinc-finger nucleases , 2010, Nature Methods.

[7]  Jin-Soo Kim,et al.  Surrogate reporters for enrichment of cells with nuclease-induced mutations , 2011, Nature Methods.

[8]  J. Orange,et al.  Establishment of HIV-1 resistance in CD4+ T cells by genome editing using zinc-finger nucleases , 2008, Nature Biotechnology.

[9]  James E. DiCarlo,et al.  RNA-Guided Human Genome Engineering via Cas9 , 2013, Science.

[10]  Feng Zhang,et al.  CRISPR-assisted editing of bacterial genomes , 2013, Nature Biotechnology.

[11]  Claudio Mussolino,et al.  A novel TALE nuclease scaffold enables high genome editing activity in combination with low toxicity , 2011, Nucleic acids research.

[12]  Jon Cohen The emerging race to cure HIV infections. , 2011, Science.

[13]  J. Doudna,et al.  RNA-guided genetic silencing systems in bacteria and archaea , 2012, Nature.

[14]  Elo Leung,et al.  A TALE nuclease architecture for efficient genome editing , 2011, Nature Biotechnology.

[15]  Eunji Kim,et al.  Precision genome engineering with programmable DNA-nicking enzymes , 2012, Genome research.

[16]  Jeffry D. Sander,et al.  Efficient In Vivo Genome Editing Using RNA-Guided Nucleases , 2013, Nature Biotechnology.

[17]  Eunji Kim,et al.  Targeted chromosomal duplications and inversions in the human genome using zinc finger nucleases. , 2012, Genome research.