Efficient targeting of expressed and silent genes in human ESCs and iPSCs using zinc-finger nucleases

被引：790
作者：
Hockemeyer, Dirk [1 ]
Soldner, Frank [1 ]
Beard, Caroline [1 ]
Gao, Qing [1 ]
Mitalipova, Maisam [1 ]
DeKelver, Russell C. [3 ]
Katibah, George E. [3 ]
Amora, Ranier [3 ]
Boydston, Elizabeth A. [3 ]
Zeitler, Bryan [3 ]
Meng, Xiangdong [3 ]
Miller, Jeffrey C. [3 ]
Zhang, Lei [3 ]
Rebar, Edward J. [3 ]
Gregory, Philip D. [3 ]
Urnov, Fyodor D. [3 ]
Jaenisch, Rudolf [1 ,2 ]
机构：
[1] Whitehead Inst Biomed Res, Cambridge, MA 02142 USA
[2] MIT, Dept Biol, Cambridge, MA USA
[3] Sangamo BioSci Inc, Richmond, CA USA
来源：
NATURE BIOTECHNOLOGY | 2009年 / 27卷 / 09期
基金：
美国国家卫生研究院;
关键词：
EMBRYONIC STEM-CELLS; HOMOLOGOUS RECOMBINATION; SOMATIC-CELLS; ELECTROPORATION; IDENTIFICATION; PLURIPOTENCY; INTEGRATION; DISEASE; LOCUS; LINE;
D O I：
10.1038/nbt.1562
中图分类号：
Q81 [生物工程学（生物技术）]; Q93 [微生物学];
学科分类号：
071005 ; 0836 ; 090102 ; 100705 ;
摘要：
Realizing the full potential of human embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs) requires efficient methods for genetic modification. However, techniques to generate cell type-specific lineage reporters, as well as reliable tools to disrupt, repair or overexpress genes by gene targeting, are inefficient at best and thus are not routinely used. Here we report the highly efficient targeting of three genes in human pluripotent cells using zinc-finger nuclease (ZFN)-mediated genome editing. First, using ZFNs specific for the OCT4 (POU5F1) locus, we generated OCT4-eGFP reporter cells to monitor the pluripotent state of hESCs. Second, we inserted a transgene into the AAVS1 locus to generate a robust drug-inducible overexpression system in hESCs. Finally, we targeted the PITX3 gene, demonstrating that ZFNs can be used to generate reporter cells by targeting non-expressed genes in hESCs and hiPSCs.
引用
收藏
页码：851 / U110
页数：9
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