绿色荧光蛋白表达质粒的构建及鉴定

目的构建能表达绿色荧光蛋白的载体———pCDNA3.1-GFP。方法使用基因重组方法从质粒pGreenlantern中获取绿色荧光蛋白基因片段,克隆入PCDNA3.1的多克隆区,构建成PCDNA3.1-GFP,通过限制性核酸内切酶NotI与BamHI对所建质粒进行分析后,转染HepG2细胞并观察绿色荧光蛋白表达。结果经限制性内切酶及转染HepG2细胞表达鉴定,证实成功构建了PCDNA3.1-GFP。结论成功构建成携有绿色荧光蛋白报告基因哺乳动物表达载体,该质粒可用于融合表达目的蛋白,并将目的蛋白表达定位。

The construction and identification of the plasmid expressing green fluorescent protein

AIM To construct the vector PCDNA3.I-GFP expressing green fluorescent protein tag. Methods The vector pGreen lantern was digested with NotI, the encoding region of green fluorescent protein tag was inserted into the MCS of the PCDNA3.1, the recombinant was confirmed by restriction enzyme map and transfected into HepG2 cells to ensure the expression of green fluorescent protein. Results The Results of the restriction enzyme digestion confirmed the recombinant vector was constructed successfully. furthermore it can express green fluorescent protein in HepG2 cells. Conclusion The vector PCDNA3.1-GFP was constructed successfully, It is convenient to orientate your interested protein through fusing expression.It also laid foundation for making the vector as a reporter gene plasmid.