Cell Migration and Invasion Group, Department of Cell Biology, UMR144, Institut Curie, Paris, France.
Institut Curie Plateforme d'Expérimentation In Vivo, Université Paris-Sud 11, Orsay, France.
Methods Mol Biol. 2021;2214:125-141. doi: 10.1007/978-1-0716-0958-3_9.
CRISPR /Cas9 is a powerful technology that has transformed gene editing of mammalian genomes, being faster and more cost-effective than standard gene targeting techniques. In this chapter, we provide a step-by-step protocol to obtain Knock-Out (KO ) or Knock-In (KI ) mouse models using CRISPR /Cas9 technology. Detailed instructions for the design of single guide RNAs (sgRNA ) for KO approaches and single-strand oligonucleotide (ssODN ) matrix for generation of KI animals are included. We also describe two independent CRISPR /Cas9 delivery methods to produce gene-edited animals starting from zygote-stage embryos, based either on cytoplasmic injection or electroporation.
CRISPR/Cas9 是一种强大的技术，它改变了哺乳动物基因组的基因编辑，比标准的基因靶向技术更快、更具成本效益。在这一章中，我们提供了一个使用 CRISPR/Cas9 技术获得敲除（KO）或敲入（KI）小鼠模型的分步方案。我们还描述了两种独立的 CRISPR/Cas9 递送方法，从受精卵阶段的胚胎开始，基于细胞质注射或电穿孔，产生基因编辑动物。
我们还提供了用于 KO 方法的单个指导 RNA（sgRNA）和用于生成 KI 动物的单链寡核苷酸（ssODN）矩阵的设计的详细说明。
