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Genetic engineering of human pluripotent cells using TALE nucleases


Targeted genetic engineering of human pluripotent cells is a prerequisite for exploiting their full potential. Such genetic manipulations can be achieved using site-specific nucleases. Here we engineered transcription activator–like effector nucleases (TALENs) for five distinct genomic loci. At all loci tested we obtained human embryonic stem cell (ESC) and induced pluripotent stem cell (iPSC) clones carrying transgenic cassettes solely at the TALEN-specified location. Our data suggest that TALENs employing the specific architectures described here mediate site-specific genome modification in human pluripotent cells with similar efficiency and precision as do zinc-finger nucleases (ZFNs).

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Figure 1: Genetic engineering of ESCs and iPSCs using TALENs.


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We thank R. Alagappan, P. Xu, S. Cristea, S. Lam and A. Vincent for expert technical assistance. We thank F. Soldner for helpful discussions on the manuscript. D.H. is a Merck Fellow of the Life Sciences Research Foundation. R.J. was supported by US National Institutes of Health grants R37-CA084198, RO1-CA087869 and RO1-HD045022 and by a grant from the Howard Hughes Medical Institute.

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Authors and Affiliations



D.H., H.W. and R.J. designed the targeting experiments and wrote the manuscript. H.W. and D.H. generated donor plasmids. D.H. preformed targeting experiments. S.K., C.S.L. and H.W. assisted with Southern blot analysis. Q.G. analyzed teratomas. J.P.C. and D.H. performed FACS analysis of targeted cells. L.Z. and J.C.M. designed the TALENs, S.J.H. assembled the TALENs, G.J.C. and Y.S. tested the TALENs, and B.Z., J.M.C. and X.M. performed the off-target analysis. D.H., R.J., L.Z., G.J.C., J.C.M., B.Z., X.M. and F.D.U. analyzed the data. E.J.R., P.D.G. and F.D.U. designed and supervised the design of the TALENs and contributed to writing the manuscript.

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Correspondence to Rudolf Jaenisch.

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Competing interests

R.J. is an adviser to Stemgent and a cofounder of Fate Therapeutics. G.J.C., L.Z., Y.S., J.C.M., B.Z., J.M.C., X.M., S.J.H., E.J.R., P.D.G. and F.D.U. are full-time employees of Sangamo BioSciences.

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Hockemeyer, D., Wang, H., Kiani, S. et al. Genetic engineering of human pluripotent cells using TALE nucleases. Nat Biotechnol 29, 731–734 (2011).

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