Gene targeting by homologous recombination in mouse zygotes mediated by zinc-finger nucleases
Gene targeting by homologous recombination in mouse zygotes mediated by zinc-finger nucleases
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DOI：
10.1073/pnas.1009424107
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发表时间：
2010-08-24
影响因子：
11.1
通讯作者：
Kuehn, Ralf
中科院分区：
文献类型：
--
作者：
Meyer, Melanie;de Angelis, Martin Hrabe;Kuehn, Ralf
在胚胎干细胞中通过同源重组进行基因打靶被广泛用于产生特定的小鼠突变体。然而，大多数哺乳动物物种缺乏用于靶向基因操作的工具。由于双链断裂极大地提高了基因组位点上同源重组的速率，我们探究了是否可以通过使用锌指核酸酶在受精卵中直接进行基因打靶。在此我们报道，在将靶向载体与锌指核酸酶共同注射到小鼠单细胞胚胎中时，在没有预筛选的情况下，1.7 - 4.5%的胚胎实现了基因打靶。这些发现使得在受精卵中一步操作哺乳动物生殖系成为可能，且不依赖于胚胎干细胞。
Gene targeting by homologous recombination in embryonic stem cells is extensively used to generate specific mouse mutants. However, most mammalian species lack tools for targeted gene manipulation. Since double-strand breaks strongly increase the rate of homologous recombination at genomic loci, we explored whether gene targeting can be directly performed in zygotes by the use of zinc-finger nucleases. Here we report that gene targeting is achieved in 1.7-4.5% of murine one-cell embryos upon the coinjection of targeting vectors with zinc-finger nucleases, without preselection. These findings enable the manipulation of the mammalian germ line in a single step in zygotes, independent of ES cells.