【摘要】 目的 构建能复合表达绿色荧光蛋白和白介素－２重组卡介菌（ｒＢＣＧ０ＧＦＰ－ＩＬ－２）。方法 采用基因 工程技术构建ｐＤｍＧＦＰ－ＩＬ－２重组穿梭质粒，用电穿孔法将重组质粒ｐＤＭＧＦＰ－ＩＬ－２转化大肠杆菌和卡介菌中，采用限制酶 切，荧光显微锐下观察及ＰＣＲ方法鉴定 ｐＤｍＧＦＰ－ＩＬ－２和 ｒＢＣＧ－ＧＦＰ－ＩＬ－２。结果 琼脂糖凝胶电泳结果显示 ｐＤｍＧＦＰ－１１．２质 粒的限制酶切条带，ＰＣＲ法检测重组ＢＣＧ菌株可见明显ＩＬ－２片段扩增条带约０．３ｋｂ，ｒＢＣＧ－ＧＦＰ－ＩＬ－２液体培养菌液在长 波紫外线的照射下，发出明亮的绿色荧光，小鼠皮下接种ｒＢＣＧ－ＧＦＰ－ＩＬ－２．１周后，取组织冰冻切片在荧光显微镜下可见 绿色荧光。结论 成功构建了重组质粒ｐＤｍＧＦＰ－ＩＬ－２和共表达绿色荧光蛋白和白介素－２重组ＢＣＧ（ｒＢＣＧ－ＧＦＰ－ＩＬ－２）。更多还原

【Abstract】 Objective To construct a recombinant BCG( rBCG-TFP-IE2) for the co-expression of green fluorescent protein and IL-2. Methods A recombinant plasmid pDmGFP-IL-2 was constructed by gene engineering technique, identified by digestion with restriction endonuclease, and transformed to E. coli and BCG by electric perforation. The expressed product was identified with fluorescence microscopy and PCR. Re-sults Agarose gel electropharesis showed that IL-2 gene was inserted into pDmGFP-IL-2. A 0.3kb of IL-2 fragment was amplified from rBCG-GFP-IL-2 by PCR. The rBCG-GFP-IL-2 liquid culture showed bright green fluorescence in the light of a long-wave ultraviolet ray. The frozen tissue section of mice one week after being subcutaneously inoculated with rBCG-GFP-IL-2 also showed green fluorescence under microscope. Conclusion Recombinant plasmid pDmGFP-IL-2 and rBCG-GFP-IL-2 were successfully constructed.更多还原