人工合成启动子特异启动IGF-1真核载体的构建与绵羊成纤维细胞转染

Construction of a Muscle Specific Expression Vector of IGF-1 Drived by Synthetic Muscle Promoter and Its Transfection into Sheep Fetal Fibroblasts Cells

构建了肌肉特异表达IGF-1载体,并转染绵羊成纤维细胞获得了稳定整合外源基因可用于细胞核移植的转基因细胞.通过人工合成骨骼肌特异启动子SP片段,与羊IGF-1相连,最终连接到骨架载体pCDsRed2中构建成骨骼肌特异表达IGF-的真核表达载体pSPICDS.使用脂质体法转染绵羊成纤维细胞,通过红色荧光与G418双重选择,获得了转基因细胞.经PCR检测证实得到的细胞克隆外源基因稳定整合,可以作为供体细胞用于体细胞核移植.

A muscle specific expression vector of ovine insulin like growth factor I （IGF-1） was constructed,then sheep fetal fibroblast cells （SFCs） were transfected by this vector,and transgenic SFCs which exogenous gene inserted into the cell genomic were obtained. The synthetic muscle promoter was connected with ovine IGF-1, then linked with the backbone vector pCdsRed2, then the vector named pSPICDS was obtained. The LipofectamineTM was used to transfect the SFCs,and the transgenic SFCs were selected with G418 and red fluorescence. The transgenic SFCs were analysed by PCR and the results demonstrated that the exogenous genes were stably integrated into the SFCs genome,and the transgenic SFCs could be used as donor cells of nuclear transfer to produce transgenic sheeps.