Genetically modified mouse models are essential for in vivo investigation of gene function and human disease research. Targeted mutations can be introduced into mouse embryos using genome editing technology such as CRISPR-Cas. Although mice with small indel mutations can be produced, the production of mice carrying large deletions or gene fragment knock-in alleles remains inefficient. We introduced the nuclear localisation property of Cdt1 protein into the CRISPR-Cas system for efficient production of genetically engineered mice. Mouse Cdt1-connected Cas9 (Cas9-mC) was present in the nucleus of HEK293T cells and mouse embryos. Cas9-mC induced a bi-allelic full deletion of Dmd, GC-rich fragment knock-in, and floxed allele knock-in with high efficiency compared to standard Cas9. These results indicate that Cas9-mC is a useful tool for producing mouse models carrying targeted mutations.

中文翻译：

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在小鼠受精卵中使用 Cas9-mouse Cdt1 进行基因组编辑介导的大缺失和基因片段敲入小鼠的高效生产

转基因小鼠模型对于基因功能的体内研究和人类疾病研究至关重要。可以使用 CRISPR-Cas 等基因组编辑技术将靶向突变引入小鼠胚胎中。虽然可以生产具有小的 indel 突变的小鼠，但携带大缺失或基因片段敲入等位基因的小鼠的生产仍然效率低下。我们将 Cdt1 蛋白的核定位特性引入 CRISPR-Cas 系统，以有效生产基因工程小鼠。小鼠 Cdt1 连接的 Cas9 (Cas9-mC) 存在于 HEK293T 细胞和小鼠胚胎的细胞核中。与标准 Cas9 相比，Cas9-mC 诱导了 Dmd 的双等位基因完全缺失、富含 GC 的片段敲入和 floxed 等位基因敲入。