肉桂地链霉菌接合转移体系的构建及nsdA基因中断对其次级代谢的影响  

Construction of the Conjugal Transfer System of Streptomyces cinnamonensis and Effect of PCR-mediated nsdA Gene Disruption on Its Secondary Metabolism

作　　者：陈芬[1] 熊伟[2] 闵勇[1] 范与庆[1] 梁运祥[1] 吕和平[3] 邢仁昌[3] 郑应华[1] 

机构地区：[1]华中农业大学农业微生物国家重点实验室,武汉430070 [2]清华大学生物科学与技术系,北京100084 [3]北京生物医药研究所,北京100091

出　　处：《农业生物技术学报》2007年第6期1042-1047,共6页Journal of Agricultural Biotechnology

摘　　要：以pSET152和pHL212为出发质粒,通过供体大肠杆菌(Escherichia coli)ET12567(pUZ8002)和S17-1属间接合转移肉桂地链霉菌(Streptomyces cinnamonensis),构建并优化了肉桂地链霉菌的接合转移系统。利用PCR介导的基因置换技术快速构建了肉桂地链霉菌nsdA(negative regulator of Streptomyces differentiation)基因中断载体,通过接合转移导入肉桂地链霉菌工业菌株BIB2005,筛选得到1株遗传稳定表型为AprRKanS的接合子BIB309。PCR分析结果显示,该接合子即为nsdA中断突变株。与出发工业菌株相比,在摇瓶水平上nsdA中断突变株BIB309莫能菌素产能提高了2.7倍。Intergeneric transfer of plasmid vectors pSET152 and pHL212 from donor Escherichia coli ET 12567 （pUZ8002） and S17-1 to Streptomyces cinnamonensis were demonstrated and optimized. The nsdA gene disruption structure was constructed by PCR-targeting system and then introduced into S. cinnamonensis BIB2005 through intergeneric conjugal transfer. After PCR analysis, the screened Apr^RKan^s conjugant BIB309 was confirmed to be the nsdA mutant. Compared with the start strain, the yield of monensin of the nsdA mutant BIB309 increased 270 percent in the level of flask.

关 键 词：肉桂地链霉菌 接合转移 nsdA基因 基因中断 

分 类 号：S188[农业科学—农业基础科学]