Gene targeting of a disease-related gene in human induced pluripotent stem and embryonic stem cells.
Gene targeting of a disease-related gene in human induced pluripotent stem and embryonic stem cells.
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DOI：
10.1016/j.stem.2009.05.023
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发表时间：
2009-07-02
期刊：
影响因子：
23.9
通讯作者：
Cheng L
中科院分区：
文献类型：
--
作者：
Zou J;Maeder ML;Mali P;Pruett-Miller SM;Thibodeau-Beganny S;Chou BK;Chen G;Ye Z;Park IH;Daley GQ;Porteus MH;Joung JK;Cheng L
我们在此报道了在人类诱导多能干细胞（iPS）和胚胎干细胞（ES）中通过同源重组（HR）介导的对两个不同基因的基因打靶。在两种细胞类型中，用表达锌指核酸酶（ZFNs）的质粒作为供体DNA转染后，通过HR介导的对染色体整合的突变绿色荧光蛋白（GFP）报告基因的校正效率达到0.14% - 0.24%。经改造的能在GFP基因中诱导序列特异性双链断裂的ZFNs使HR介导的校正增强了>1400倍，且在干细胞核型或多能性方面未检测到改变。在内源PIG - A位点也实现了高效的HR介导的插入诱变，针对该基因的ZFNs使其增强了>200倍。容易获得具有正常核型的PIG - A基因缺失的人类ES和iPS细胞克隆。PIG - A转基因表达可挽救表型和生物学缺陷。我们的研究首次证明了在人类iPS细胞中HR介导的基因打靶，以及ZFNs在人类iPS细胞和ES细胞中诱导特定基因修饰的能力。
We report here homologous recombination (HR)-mediated gene targeting of two different genes in human iPS and ES cells. HR-mediated correction of a chromosomally-integrated mutant GFP reporter gene reaches efficiencies of 0.14%-0.24% in both cell types transfected by donor DNA with plasmids expressing zinc-finger nucleases (ZFNs). Engineered ZFNs that induce a sequence-specific double-strand-break in the GFP gene enhanced HR-mediated correction by >1400-fold without detectable alterations in stem cell karyotypes or pluripotency. Efficient HR-mediated insertional mutagenesis was also achieved at the endogenous PIG-A locus, with a >200-fold enhancement by ZFNs targeted to that gene. Clonal PIG-A null human ES and iPS cells with normal karyotypes were readily obtained. The phenotypic and biological defects were rescued by PIG-A transgene expression. Our study provides the first demonstration of HR-mediated gene targeting in human iPS cells, and the power of ZFNs for inducing specific genetic modifications in human iPS as well as ES cells.
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影响因子：
4.5
作者：
Pruett-Miller SM;Reading DW;Porter SN;Porteus MH
通讯作者：
Porteus MH
影响因子：
64.8
作者：
Park, In-Hyun;Zhao, Rui;Daley, George Q.
通讯作者：
Daley, George Q.
影响因子：
12.4
作者：
Pruett-Miller, Shondra M.;Connelly, Jon P.;Porteus, Matthew H.
通讯作者：
Porteus, Matthew H.
DOI：
10.1073/pnas.0806976105
发表时间：
2008-09-16
影响因子：
11.1
作者：
Suzuki, Keiichiro;Mitsui, Kaoru;Mitani, Kohnosuke
通讯作者：
Mitani, Kohnosuke
影响因子：
44.1
作者：
Cai, Liuhong;Ye, Zhaohui;Cheng, Linzhao
通讯作者：
Cheng, Linzhao