Department of Physiology, School of Medicine, Keio University, Shinjuku-ku, Tokyo, 160-8582, Japan.
Laboratory for Proteolytic Neuroscience, RIKEN Center for Brain Science, Wako City, Saitama, 351-0198, Japan.
Sci Rep. 2019 Feb 6;9(1):1528. doi: 10.1038/s41598-018-37990-w.
Genome editing technology greatly facilitates the genetic modification of various cells and animals. The common marmoset (Callithrix jacchus), a small non-human primate which exhibits high reproductive efficiency, is a widely used animal model in biomedical research. Developing genome editing techniques in the common marmoset will further enhance its utility. Here, we report the successful establishment of a knock-in (KI) method for marmoset embryonic stem cells (ESCs), which is based on the CRISPR-Cas9 system. The use of CRISPR-Cas9, mediated by homologous recombination (HR), enhanced the KI efficiency in marmoset ESCs. Furthermore, we succeeded in performing KI in early-stage marmoset embryos. In the course of the experiments, we found that HR in the marmoset ESCs is innately highly efficient. This suggested that the marmoset possesses a repair mechanism for DNA double-strand breaks. The current study will facilitate the generation of genetically modified marmosets and gene function analysis in the marmoset.
基因组编辑技术极大地促进了各种细胞和动物的基因修饰。普通狨猴（Callithrix jacchus）是一种小型非人类灵长类动物，具有较高的繁殖效率，是生物医学研究中广泛使用的动物模型。开发普通狨猴的基因组编辑技术将进一步增强其效用。在这里，我们报告了一种基于 CRISPR-Cas9 系统的普通狨猴胚胎干细胞（ESC）基因敲入（KI）方法的成功建立。利用同源重组（HR）介导的 CRISPR-Cas9 提高了普通狨猴 ESC 的 KI 效率。此外，我们还成功地在早期普通狨猴胚胎中进行了 KI。在实验过程中，我们发现普通狨猴 ESC 中的 HR 具有先天的高效率。这表明普通狨猴具有一种修复 DNA 双链断裂的机制。本研究将有助于生成基因修饰的普通狨猴和普通狨猴中的基因功能分析。
