蛋白A-绿色荧光蛋白融合蛋白的构建与表达

绿色荧光蛋白 (GFP)是一种生物发光蛋白。GFP作为基因表达标记物或融合蛋白标记在动物、植物、微生物的研究中广泛应用。本实验将gfp基因与蛋白A基因重组 ,在大肠杆菌中融合表达为A GFP蛋白。根据gfp基因与质粒载体pRIT2T的序列 ,设计一对引物 ,以PCR技术扩增gfp基因序列 ,克隆到表达载体pRIT2T的EcoRⅠ与BamHⅠ酶切位点 ,与载体中的A蛋白基因融合。重组质粒转化大肠杆菌Top10 ,菌落在 36 5nm紫外光下呈现明亮的绿色荧光。本研究成功构建并表达了GFP与蛋白A的重组质粒 ,为开展GFP的研究及应用提供了生物工程工具的前体。

Expression of fused protein A-green fluorescent protein (PA-GFP)

The green fluorescent protein (GFP) of jellyfish victoria is an unusual protein with strong visible fluorescence both in vivo and in vitro. In this study, about 750 bp fragment of gfp gene was amplified by PCR and was inserted into pRIT2T vector containing protein A gene. The recombinant plasmid was transferred into E.coli JM101. GFP and protein A were fused and expressed in E.coli Top10. The bright green fluorescence on the plates could be visible under UV at 365 nm. The bacteria cells were disrupted by sonic oscillation. The fused protein PA|GFP could be obtained from the supernatants.