Journal Article

Efficient generation of Knock-in/Knock-out marmoset embryo via CRISPR/Cas9 gene editing

Sato, Kenya, Sato, Kengo, Kurotaki, Yoko, Kishi, Noriyuki, Harada, Takeshi, Aiba, Atsu, Sakakibara, Yasubumi, Kumita, Wakako, Feng, Guoping, Suzuki, Yasuhiro, Zhou, Yang, Sasaki, Erika, Okano, Hideyuki

2019

Overview

Genetically modified nonhuman primates (NHP) are useful models for biomedical research. Gene editing technologies have enabled production of target-gene knock-out (KO) NHP models. Target-gene-KO/knock-in (KI) efficiency of CRISPR/Cas9 has not been extensively investigated in marmosets. In this study, optimum conditions for target gene modification efficacies of CRISPR/mRNA and CRISPR/nuclease in marmoset embryos were examined. CRISPR/nuclease was more effective than CRISPR/mRNA in avoiding mosaic genetic alteration. Furthermore, optimal conditions to generate KI marmoset embryos were investigated using CRISPR/Cas9 and 2 different lengths (36 nt and 100 nt) each of a sense or anti-sense single-strand oligonucleotide (ssODN). KIs were observed when CRISPR/nuclease and 36 nt sense or anti-sense ssODNs were injected into embryos. All embryos exhibited mosaic mutations with KI and KO, or imprecise KI, of c-kit . Although further improvement of KI strategies is required, these results indicated that CRISPR/Cas9 may be utilized to produce KO/KI marmosets via gene editing.

• Publisher: Nature Publishing Group UK,Nature Publishing Group
• Subject: 631/1647/1513/1967/3196 / 631/61/17 / Animals / Animals, Genetically Modified - genetics / Antisense oligonucleotides / c-Kit protein / Callithrix / CRISPR / CRISPR-Cas Systems / Embryo, Mammalian / Embryos / Gene Editing / Gene Knock-In Techniques / Gene Knockout Techniques / Genome editing / Humanities and Social Sciences / mRNA / multidisciplinary / Nuclease / RNA modification / Science / Science (multidisciplinary)