苏云金芽胞杆菌bkd基因簇的转录调控  

Transcriptional regulation of bkd gene cluster in Bacillus thuringiensis

作　　者：王冠男[1,2] 彭琦[2] 郑庆云[1,2] 李杰[1] 张杰[2] 宋福平[2] 

机构地区：[1]东北农业大学生命科学学院,黑龙江哈尔滨150030 [2]中国农业科学院植物保护研究所植物病虫害生物学国家重点实验室,北京100193

出　　处：《微生物学报》2014年第10期1129-1137,共9页Acta Microbiologica Sinica

基　　金：国家自然科学基金项目(31270111)~~

摘　　要：【目的】通过分析苏云金芽胞杆菌bkd基因簇的转录调控和bkdR突变体的表型特征,明确bkdR所在基因簇的转录调控机制和对Cry蛋白产量的影响。【方法】通过生物信息学方法分析bkdR所在基因簇的结构,RT-PCR分析基因簇的转录单元,采用同源重组技术敲除苏云金芽胞杆菌HD73菌株的bkdR基因,利用启动子融合lacZ的方法分析启动子的转录活性。利用总蛋白定量确定Cry1Ac蛋白产量。【结果】bkd基因簇由8个基因组成,其中ptb-bkdB7个基因组成1个转录单元。ptb基因的启动子转录活性在sigL和bkdR突变体中均明显降低。bkdR基因的缺失对菌体生长、芽胞形成率和Cry1Ac蛋白产量无影响,但使运动能力减弱。【结论】bkd操纵子受Sigma 54控制,并由BkdR激活,bkdR基因的缺失对Cry蛋白产量无影响,对菌株的运动能力有影响。[Objective] In order to determine the effect of bkdR deletion on Cry protein production. We analyzed the transcriptional regulation of bkd gene cluster and the phenotype of bkdR mutant. [Methods]Sequence of bkd gene cluster in Bacillus thuringiensis was analyzed by sequence alignment. RT-PCRwas used to reveal the transcriptional units of the bkd gene cluster. bkdR insertion mutant was constructed by homologous recombination. Transcriptional activity was analyzed by promoter fusions with lacZ gene. Comparison of the Cry1 Ac protein production was determined by protein quantitation. [Results]The bkd gene cluster was composed of eight genes. The ptb-bkdB formed one transcriptional unit.The transcriptional activity of ptb sharply decreased in sigL and bkdR mutants. Deletion of bkdR decreased the motility of cells,but no effect on growth,sporulation efficiency and Cry protein production. The bkd gene cluster is controlled by Sigma 54 and activated by BkdR. Deletion of bkdR has no effect on Cry protein production,but decreased the motility of the cells. [Conclusion]The bkd gene cluster is controlled by Sigma 54 and activated by BkdR. Deletion of bkdR has no effect on Cry protein production,but decreased the motility of the cells. It suggested that deletion of bkdR do not affect the Cry protein production the same as sigL mutant. It means decreasing of Cry protein productioninsigL mutant was not caused by only one EBP mutation,but might be multiple roles.

关 键 词：苏云金芽胞杆菌 增强子结合蛋白(Enhancer BINDING Proteins 简称EBPs) bkdR基因 转录起始因子Sigma54 

分 类 号：Q933[生物学—微生物学]