动物基因组定点整合转基因技术研究进展

摘要

The traditional transgenic technologies,such as embryo microinjection,transposon-mediated integration,or lentiviral transfection,usually result in random insertions of the foreign DNA into the host genome,which could have various disadvantages in the establishment of transgenic animals.Therefore,a strategy for site-specific integration of a transgene is needed to generate genetically modified animals with accurate and identical genotypes.However,the efficiency for site-specific integration of transgene is very low,which is mainly caused by two issues.The first one is the low efficiency of inducing double-strand break (DSB) at the target site of host genome in the initial process.The second one is the low efficiency of homologous recombination repair (HDR) between the target site and the donor plasmid carrying homologous arm and foreign genes.HDR is the most common mechanism for site-specific integration of a transgene.DSBs can stimulate DNA repair mainly by two competitive mechanisms,HDR and nonhomologous end joining (NHEJ).Hence,activation of HDR or inhibition of NHEJ can promote the HDR in the integration processes,thereby optimizing a specific targeting of the transgene.In this review,we summarize the recent advances in strategies for improving the site-specific integration of foreign transgene in transgenic technologies.%传统转基因技术,如显微注射、转座子、慢病毒转染等将目的基因插入基因组内的整合方式是随机的,这些随机整合对后期转基因动物品系组建和育种带来诸多不利,因此有研究人员提出了定点整合转基因技术.目前该技术的定点整合效率非常低,主要取决于两个方面:一是靶位点产生DNA双链断裂(double-strand break,DSB)的效率;二是断裂后的靶位点与携带同源臂及外源基因的供体质粒发生同源重组的效率,其中同源重组修复(homologous recombination repair,HDR)是基因组定点整合最为依赖的修复机制.靶位点产生DSB后,机体的DNA修复既可能发生HDR,也可能发生非同源末端连接(nonhomologous end joining,NHEJ),并且两者之间存在竞争关系,因此激活HDR或抑制NEHJ都可提高定点整合转基因的效率.本文结合影响定点整合的因素,对提高定点整合效率最新探索方面进行了综述.