Department of Molecular Life Science, Division of Basic Medical Science and Molecular Medicine, School of Medicine, Tokai University, Isehara, Kanagawa, Japan.
Center for Matrix Biology and Medicine, Graduate School of Medicine, Tokai University, Isehara, Kanagawa, Japan.
Genome Biol. 2018 Feb 26;19(1):25. doi: 10.1186/s13059-018-1400-x.
We present a robust method called improved-Genome editing via Oviductal Nucleic Acids Delivery (i-GONAD) that delivers CRISPR ribonucleoproteins to E0.7 embryos via in situ electroporation. The method generates mouse models containing single-base changes, kilobase-sized deletions, and knock-ins. The efficiency of i-GONAD is comparable to that of traditional microinjection methods, which rely on ex vivo handling of zygotes and require recipient animals for embryo transfer. In contrast, i-GONAD avoids these technically difficult steps, and it can be performed at any laboratory with simple equipment and technical expertise. Further, i-GONAD-treated females retain reproductive function, suggesting future use of the method for germline gene therapy.
我们提出了一种名为通过输卵管核酸递送改良基因组编辑（i-GONAD）的稳健方法，该方法通过原位电穿孔将 CRISPR 核糖核蛋白递送至 E0.7 胚胎。该方法生成了含有单碱基变化、千碱基大小缺失和基因敲入的小鼠模型。i-GONAD 的效率可与传统的微注射方法相媲美，后者依赖于对受精卵的体外处理，并且需要受体动物进行胚胎移植。相比之下，i-GONAD 避免了这些技术上困难的步骤，并且可以在任何具有简单设备和技术专长的实验室中进行。此外，经过 i-GONAD 处理的雌性保留了生殖功能，这表明该方法未来可用于生殖细胞基因治疗。
