摘要
目的 构建绿色荧光蛋白融合表达HBsAg基因的载体 ,并检测其在COS 7细胞中的表达。方法 采用PCR获得HBsAg基因 ;利用该基因片段和质粒pEGFP 3.1的HindⅢ /KpnⅠ限制性酶切位点构建融合表达载体pGHBs 3.1;通过脂质体介导的方法将该载体转染到COS 7细胞中 ,用PCR法检测基因转染 ,ELISA检测HBsAg的瞬时表达 ,荧光显微镜检测绿色荧光蛋白表达。结果 融合表达载体pGHBs 3.1转染COS 7细胞后 ,在细胞中检测到HBsAg基因片段 ,转染细胞培养上清中检测到HBsAg表达 ,用荧光显微镜观察到转染细胞中有绿色荧光蛋白表达。结论 表达载体pGHBs 3.1在COS 7细胞中获得了表达 ,表达的融合蛋白具有HBsAg和绿色荧光蛋白的双重活性 ,该载体可用绿色荧光蛋白作为报告基因观察HBsAg的表达及定位。
Objective To construct a HBsAg and EGFP fusion gene and express the gene in COS 7 cells.Methods Amplify HBsAg gene by PCR and inster the gene into plasmid pEGF P 3 1.Transfect CDS 7 cells with the recombinant plasmid pGHBs 3.1,detect the expressed HBsAg by ELISA and observe the expressed GFP under fluorescence microscope.Results HBsAg gene was detected in the transfected COS 7 cells.The expressed HBsAg was detected in the culture supernatant of the transfected cells,meanwhile the expressed GFP was observed under fluorescence microscope.Conclusion The HBsAg and EGFP fusion gene was expressed in COS 7 cells.The expressed fusion protein showed double activity of HBsAg and GFP.GFP can be used as a reported gene for tracing the expression HBsAg gene.
出处
《中国生物制品学杂志》
CAS
CSCD
2004年第2期68-70,共3页
Chinese Journal of Biologicals
