【摘要】 采用PCR技术,亚克隆来自短小芽孢杆菌总基因组中的启动子活性片段F1,构建了一个大肠杆菌 短小芽孢杆菌穿梭表达载体pHY300 F1gfp,以缺失启动子的绿色荧光蛋白(GFP)基因为报告基因,检测了该片段在短小芽孢杆菌DX01菌株中的启动活性,在荧光显微镜下,观察到了明亮的绿色荧光,证实活性片段F1具有组成型启动子的功能,GFP基因在短小芽孢杆菌中实现了组成型表达.更多还原

【Abstract】 The green fluorescent protein(GFP) of the jellyfish Aequorea Victoria is a useful reporter molecule for monitoring gene expression in vivo in eukaryotic and prokaryotic cells. A GFP vector was constructed for in situ detection of the rice epiphyte Bacillus brevis strain DX01. The gfpS65T gene was transcriptionally fused to a strong B. brevis promoter functional fragment F1,which was subcloned from the genome of strain DX01 by PCR technique, and then inserted into an Escherichia coliBacillus shuttle vector pHY300PLK. DX01 cells harboring pHY300F1gfp could produce bright green fluorescence when they were examined by using fluorescent microscopy. The results were confirmed by western blotting and fluorescenceactivated cell sorting (FACS). The promoter Functional fragment F1 showed constitutive activity.更多还原