piggyBac转座子在金鱼及泥鳅转基因中的应用  

Exploration of transgenic method for goldfish and loach with the piggyBac transposon

作　　者：胡莹莹[1] 郭学双[1] 周阳[1] 曹广力[1] 贡成良[1] 薛仁宇[1] 

机构地区：[1]苏州大学基础医学与生物科学学院,江苏苏州215123

出　　处：《水产学报》2012年第10期1473-1481,共9页Journal of Fisheries of China

基　　金：国家重大基础研究项目(2012CB114605)

摘　　要：为了探讨piggyBac转座子在鲤科鱼类及鳅科鱼类转基因中的适用性,实验通过将PAβ启动子控制的DsRed表达元件插入pigA3GFP中,构建转基因载体pigA3GFP-AβDsRed。用该载体对草鱼肾脏(CIK)细胞转染,结果显示,通过piggyBac转座子可将外源基因导入到CIK细胞的基因组中,来源于家蚕的A3启动子和来源于巨细胞病毒的CMV启动子在CIK细胞均具有活性。将转基因载体pigA3GFP、pigA3GFP-AβDsRed分别与辅助质粒helper-pigA3混合后以精子介导法导入金鱼和泥鳅的受精卵后,经荧光观察、PCR鉴定、Dot blotting鉴定,证实获得了转基因金鱼和转基因泥鳅。研究表明,PB转座子在鲤科鱼类及鳅科的一些鱼中具有转座活性。Development of transgenic fish techniques and application will promote traits improvement of fish and benefit exploration of its gene function. To investigate the applicability of the transposon piggyBac in Cyprinidae and Cobitidae fish, the expression element of DsRed under the control of PAp promoter was inserted into the plasmid pigA3GFP to generate transgenic vector pigA3GFP-A[3DsRed. The results of the CIK cell transfected with pigA3GFP-AI3DsRed showed that the transposon piggyBac could transfer exogenous genes into CIK genome, and both of the promoter A3 from silkworm and the promoter CMV from CMV have the bioactivity. Moreover, the pigA3GFP-A[3DsRed and pigA3GFP were mixed with helper plasmid helper-pigA3 respectively,and then were introduced into the eggs of goldfish and loach by sperm- mediated transfer. The results of fluorescence observation,PCR and Dot blotting verified that both transgenic goldfish and loach were obtained. Therefore, PB transposon was considered to be active in some fishes of Cyprinidae and Cobitidae.

关 键 词：金鱼 泥鳅 PIGGYBAC转座子 转基因 草鱼肾脏(CIK) 

分 类 号：Q782[生物学—分子生物学] S917.4[农业科学—水产科学]