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Enhanced efficiency of human pluripotent stem cell genome editing through replacing TALENs with CRISPRs.
Ding Q, Regan SN, Xia Y, Oostrom LA, Cowan CA, Musunuru K
Cell Stem Cell. 2013 Apr 4;12(4):393-4. doi: 10.1016/j.stem.2013.03.006.
PubMed Article

Kiran Musunuru’s lab has developed a human codon-optimized Cas9 nuclease from Streptococcus pyogenes, which is optimized for expression in human pluripotent stem cells and co-expresses enhanced green fluorescent protein (EGFP) for cell selection. These plasmids can be used in any human or mammalian cell and can be used in combination with the gRNA vector (Addgene ID 41824) reported in the publication below.

The Cas9 nuclease is based on the version described in:

RNA-Guided Human Genome Engineering via Cas9. Mali P, Yang L, Esvelt KM, Aach J, Guell M, Dicarlo JE, Norville JE, Church GM. Science . 2013 Feb 15;339(6121):823-6. PubMed

Resources

A detailed protocol for the use of these plasmids to perform genome editing of cells is available at StemBook: www.stembook.org/node/1438y. A PDF of the protocol can be downloaded from that page.

Plasmids from Article

ID Plasmid Purpose  
44719pCas9_GFPCo-expression of human codon-optimized Cas9 nuclease and GFP, plasmid optimized for expression in human pluripotent stem cells
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44720pCas9D10A_GFPCo-expression of human codon-optimized Cas9 (D10A) mutant nickase and GFP, plasmid optimized for expression in human pluripotent stem cells
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