通过导入表达SgRNA/Cas9的质粒对小鼠受精卵和胚胎干细胞进行基因组编辑。
Genome Editing in Mouse Zygotes and Embryonic Stem Cells by Introducing SgRNA/Cas9 Expressing Plasmids.
作者信息
Noda Taichi, Oji Asami, Ikawa Masahito
机构信息
Osaka University, 3-1 Yamada-oka, Suita, Osaka, 5650871, Japan.
Research Fellow of Japan Society for the Promotion of Science (JSPS), Tokyo, 1020083, Japan.
出版信息
Methods Mol Biol. 2017;1630:67-80. doi: 10.1007/978-1-4939-7128-2_6.
In mammalian cells, genome editing with the single guide RNA (sgRNA)/Cas9 complex allows for high targeting efficiency within a relatively short time frame with the added benefits of being low cost and easy to design. sgRNA/Cas9-mediated editing in mouse zygotes has accelerated the analysis of gene functions and the generation of mouse models of human diseases. Despite the benefits, this method still suffers from several problems, such as mosaicism in the founder generation which complicates genotyping and phenotypical analyses, and the low efficiency of more complicated genome editing. Thus, we recently established the system for genome editing in embryonic stem (ES) cells and its application for chimeric analysis in mice. In this section, we introduce the procedure for genome editing in mouse zygotes and ES cells.
在哺乳动物细胞中，利用单向导RNA（sgRNA）/Cas9复合物进行基因组编辑，能够在相对较短的时间内实现高靶向效率，而且成本低、易于设计。在小鼠受精卵中进行sgRNA/Cas9介导的编辑，加速了基因功能分析以及人类疾病小鼠模型的建立。尽管有这些优点，但该方法仍存在一些问题，如奠基者一代中的嵌合体现象，这使得基因分型和表型分析变得复杂，以及更复杂的基因组编辑效率较低。因此，我们最近建立了胚胎干细胞（ES细胞）基因组编辑系统及其在小鼠嵌合分析中的应用。在本节中，我们介绍小鼠受精卵和ES细胞基因组编辑的步骤。
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