PiggyBac转座元件的构建及其在团头鲂基因组中的转基因效率  

The study on transgenic efficiency of PiggyBac transposon in the genome of Megalobrama amblycephala

作　　者：王瑶[1] 蒋霞云[1] 邹曙明[1] 

机构地区：[1]上海海洋大学农业部淡水水产种质资源重点实验室,上海201306

出　　处：《上海海洋大学学报》2014年第2期161-166,共6页Journal of Shanghai Ocean University

基　　金：国家自然科学基金(31272633;31201760);"十二五"国家科技支撑计划(2012BAD26B00);上海高校知识服务平台(ZF1206)

摘　　要：鳞翅目昆虫粉纹夜蛾(Trichoplusia ni)的PiggyBac(PB)转座子已用于模式生物小鼠的转基因及插入诱变研究,目前,该转座子在养殖鱼类中的转基因效率如何还不清楚。构建了带PiggyBac转座子左臂、右臂、EF1α启动子和绿色荧光蛋白(eGFP)编码框的pPBs-EF1α-eGFP供体质粒。以50 ng/μL供体质粒和100 ng/μL体外转录的PB转座酶mRNA共同显微注射入团头鲂(Megalobrama amblycephala)1~2细胞期受精卵中,团头鲂eGFP的荧光表达率可达58.26%,PCR检测结果显示,该转座系统在团头鲂成鱼基因组中的整合效率为53.04%。表明PiggyBac转座子可高效介导基因在团头鲂基因组中的插入,为进一步开展团头鲂插入诱变研究奠定了基础。PiggyBac （PB） transposon is derived from Trichoplusia ni of lepidopteran and has been widely used in transgenic and insertion mutagenesis studies in mouse. Currently, relevant research about PiggyBac transposons used in farmed fish has not yet been reported. To study insertion efficiency of PiggyBac transposon in the genome of M. amblycephala, we built pPBs-EFI^-eGFP donor plasmid with PiggyBac transposon left and right arms, EFlc~ promoter and eGFP gene, then co-injected with PiggyBac transposase mRNA into the 1 -2 cell stage fertilized eggs of M. amblycephala. The concentrations of donor plasmid and transposase mRNA were 50 ng/μL and 100 ng/μL respectively. The eGFP fluorescence expression rate was 58.26%. In adult fish, PCR results demonstrated that integration efficiency of PiggyBac transposition system was 53.04% in M. amblycephala genome. Our data suggest that PiggyBac transposon can efficiently mediate gene insertion in M. amblycephala, which could be used in insertional mutagenesis in M. amblycephala.

关 键 词：PIGGYBAC转座子 团头鲂 转基因 

分 类 号：S917[农业科学—水产科学]