Wang Yong, Du Yinan, Shen Bin, Zhou Xiaoyang, Li Jian, Liu Yu, Wang Jianying, Zhou Jiankui, Hu Bian, Kang Nannan, Gao Jimin, Yu Liqing, Huang Xingxu, Wei Hong
Department of Laboratory Animal Science, College of Basic Medical Sciences, Third Military Medical University, Chongqing 400038, China.
MOE Key Laboratory of Model Animal for Disease Study, Model Animal Research Center of Nanjing University, National Resource Center for Mutant Mice, Nanjing 210061, China.
Sci Rep. 2015 Feb 5;5:8256. doi: 10.1038/srep08256.
Co-injection of zygotes with Cas9 mRNA and sgRNA has been proven to be an efficient gene-editing strategy for genome modification of different species. Genetic engineering in pigs holds a great promise in biomedical research. By co-injection of one-cell stage embryos with Cas9 mRNA and Npc1l1 sgRNA, we achieved precise Npc1l1 targeting in Chinese Bama miniature pigs at the efficiency as high as 100%. Meanwhile, we carefully analyzed the Npc1l1 sgRNA:Cas9-mediated on- and off-target mutations in various somatic tissues and ovaries, and demonstrated that injection of zygotes with Cas9 mRNA and sgRNA is an efficient and reliable approach for generation of gene-modified pigs.
将Cas9 mRNA和sgRNA共同注射到受精卵中已被证明是一种用于不同物种基因组修饰的高效基因编辑策略。猪的基因工程在生物医学研究中具有巨大潜力。通过将单细胞期胚胎与Cas9 mRNA和Npc1l1 sgRNA共同注射，我们在中国巴马小型猪中实现了对Npc1l1的精确靶向，效率高达100%。同时，我们仔细分析了Npc1l1 sgRNA:Cas9介导的各种体细胞组织和卵巢中的靶向和脱靶突变，并证明将Cas9 mRNA和sgRNA注射到受精卵中是生成基因编辑猪的一种高效且可靠的方法。
