Generation of TALEN-mediated GRdim knock-in rats by homologous recombination.

Generation of TALEN-mediated GRdim knock-in rats by homologous recombination.
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DOI:
10.1371/journal.pone.0088146
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发表时间:
2014
期刊:
影响因子:
3.7
通讯作者:
Hummler E
Hummler E
中科院分区:
综合性期刊3区
文献类型:
--
作者:
Ponce de León V;Mérillat AM;Tesson L;Anegón I;Hummler E

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转录激活因子样效应物核酸酶(TALEN)是对实验动物进行精确基因组工程改造的潜在工具。我们报道了首次在大鼠中利用转录激活因子样效应物核酸酶(TALEN)通过同源重组(HDR)进行靶向基因组整合。我们组装了TALEN,并设计了一个线性供体插入片段,其靶向大鼠糖皮质激素受体(Nr3c1)中的pA476T突变(即GRdim,该突变可阻止小鼠中的受体同源二聚化)。将TALEN mRNA和线性双链供体显微注射到大鼠单细胞胚胎中。总体而言,我们在17%的后代中观察到了靶向基因组修饰,这表明TALEN在大鼠受精卵中的切割效率很高。
Transcription Activator-Like Effector Nucleases (TALEN) are potential tools for precise genome engineering of laboratory animals. We report the first targeted genomic integration in the rat using TALENs (Transcription Activator-Like Effector Nucleases) by homology-derived recombination (HDR). We assembled TALENs and designed a linear donor insert targeting a pA476T mutation in the rat Glucocorticoid Receptor (Nr3c1) namely GRdim, that prevents receptor homodimerization in the mouse. TALEN mRNA and linear double-stranded donor were microinjected into rat one-cell embryos. Overall, we observed targeted genomic modifications in 17% of the offspring, indicating high TALEN cutting efficiency in rat zygotes.
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