用于废水处理系统中目标酵母动态解析的绿色荧光蛋白基因质粒构建

Construction of Plasmid with Green Fluorescent Protein Gene for Analysis of Yeasts in Yeast Wastewater Treatment

绿色荧光蛋白 (greenfluorescentprotein ,GFP)可用于研究复合微生物体系中特定目标功能菌的特性及动态变化 ,本研究为确立用于解析酵母细胞在混合酵母废水处理系统中的动态特性的GFP技术体系奠定了基础 .将gfp基因克隆到酵母载体pACT URA3中 ,再用构建的重组质粒转化宿主菌大肠杆菌 (EscherichiacoliJM10 9) ,扩增得到含gfp的重组质粒 ,荧光显微镜照片显示 gfp基因在大肠杆菌内得到了表达 ,但表达程度不高 ;电泳图谱及聚合酶链反应结果表明 ,含 gfp基因的质粒不是以游离的形式存在 ,而可能是以某种特殊的形式和细胞染色体发生了相互作用 .

Green fluorescent protein(GFP) can be utilized in analysis of the characteristics and distribution of a targeted strain in microbial communities. This study is the first step to establish a dynamic yeast monitoring technique in a wastewater treatment system using yeast by constructing a fluorescent yeast containing gfp gene. The gfp gene was inserted into pACT1-URA3, a powerful plasmid for introducing a foreign gene into Candida boidinii, and then transformed into E coli JM109. The gfp gene was expressed, though not very highly. The results of the electrophoresis and polymerase chain reaction suggested that the newly constructed plasmid containing gfp gene might not exist in free form in the cells, but in some special way such as interaction with the chromosome.