TGF-β_1基因真核表达载体的构建及在猪脐静脉内皮细胞中的表达  

Construction of Eukaryotic Expression Plasmid of TGF-β_1 Gene and Its Expression in Swine Umbilical Vein Endothelial Cells

作　　者：王文秀[1] 邓文[1] 张彦明[1] 代晨[1] 熊奎州[1] 谢林红[1] 温元鹏[1] 

机构地区：[1]西北农林科技大学动物医学院,杨凌712100

出　　处：《畜牧兽医学报》2008年第6期832-836,共5页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家自然科学基金项目(30471290)

摘　　要：应用RT-PCR技术从猪外周血单个核细胞扩增转化生长因子β1(Transforming growth factor-β1,TGF-β1)全基因,构建含有TGF-β1基因及EGFP报告基因的真核表达质粒pEGFP-C1-TGF-β1。采用脂质体法转染体外培养的猪脐静脉内皮细胞(SUVECs)后,通过直接荧光观察pEGFP-C1-TGF-β1融合蛋白在细胞中的分布定位,并通过RT-PCR、间接免疫荧光方法检测TGF-β1基因在SUVECs中的表达。结果在转染后1周观察到绿色荧光,RT-PCR、间接免疫荧光法检测TGF-β1表达均为阳性。本研究成功构建了含有绿色荧光蛋白基因的真核表达载体pEGFP-C1-TGF-β1,且TGF-β1基因在SUVECs中获得表达。In this study, transforming growth factor-β1 （TGF-β1） full length cDNA was cloned from swine peripheral blood mononuclear cells by RT-PCR ,the eukaryotic expression plasmid of TGF-β1 gene containing green fluorescent protein （GFP） report gene was constructed. The recombinant plasmid was transfected by lipofectin to prepared swine umbilical vein vascular endothelial cells（SUVECs）. The fluorescence expression was directly detected with fluorescence microscope, and the expression of TGF-β1 were tested by RT-PCR and indirect immunofluorescence assay （IFA）, respectively. One week after transfection,green fluorescent can be seen by fluorescent microscope;RT-PCR and indirect immunofluorescence assay showed that the expression of TGF-β1 are positive. The results indicated the successful construction of the eukaryotic expression plasmid pEGFP-C1-TGF-β1 , and showed that TGF-β1 gene was expressed efficiently in transfected SUVECs.

关 键 词：脐静脉内皮细胞 转化生长因子β1(TGF-β1) 转染 

分 类 号：S813.3[农业科学—畜牧学] Q786[农业科学—畜牧兽医]