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基因工程猪可作为动物模型研究人类疾病,也是异种器官移植最合适的供体.CRISPR/Cas9系统是高效特异的基因编辑工具.因此本文通过显微注射的方式将sg RNA和Cas9 m RNA注射入单细胞期的孤雌胚胎中,待其发育为囊胚后检测打靶效率.T7EN1酶切和TA克隆测序结果表明,本研究可以在猪的孤雌胚胎中同时实现3个基因(B2M,CIITA,GGTA1或者LDLR,LEP,LEPR)的高效敲除(敲除率分别为62.5%和50%),为使用CRISPR/Cas9系统高效、快速和经济地建立多基因修饰的猪模型提供了基础.
Genetically engineered pigs can be used as animal models to study human diseases and are the most suitable donors for xenotransplantation.CRISPR / Cas9 system is a highly efficient and specific gene editing tool.Therefore, microinjection of sg RNA and Cas9 m RNA into The single cell stage parthenogenetic embryos were tested for their target efficiency after they were developed into blastocysts.The results of T7EN1 digestion and TA cloning showed that three genes (B2M, CIITA, GGTA1 or LDLR, LEP, LEPR) knockdown rates of 62.5% and 50%, respectively, provided the basis for the efficient, rapid and economical establishment of a polygenic modified porcine model using the CRISPR / Cas9 system.