子囊霉素产生菌吸水链霉菌FIM260840基因转移系统的建立  

Construction of the Gene Transfer System for Ascomycin Producing Strain Streptomyces hygroscopicus FIM260840

作　　者：方志锴[1] 赵薇[1] 彭飞[1] 连云阳[1] 

机构地区：[1]福建省微生物研究所,福建省新药(微生物)筛选重点实验室,福建福州350007

出　　处：《生物技术》2014年第5期44-48,共5页Biotechnology

基　　金：福建省自然科学基金项目("FK506生物合成途径改造发掘含氮大环内酯类新衍生物";No.2012J01307);国家"重大新药创制"课题("创新微生物药物高效筛选与发现技术平台研究--稀有放线菌及海洋放线菌药物筛选";No.2012ZX09301002-003);福州市科技计划重点项目("福州市生物制药行业技术创新中心建设";No.2013-PT-42)资助

摘　　要：目的:建立子囊霉素产生菌吸水链霉菌FIM260840的接合基因转移体系,以便基因敲除和外源基因表达等遗传操作。方法:以整合型质粒p SET152为出发质粒,通过接合转移构建并子囊霉素产生菌FIM260840的基因转移系统。结果:12.5μg/m L安普霉素可有效筛选接合子。经PCR验证,质粒成功整合到菌株FIM260840基因组DNA中,所获接合子的安普霉素抗性高达400μg/m L以上。接合子经多次传代后,导入的质粒p SET152仍稳定整合于接合子基因组DNA上。结论:建立了高效、简便的吸水链霉菌FIM260840的基因转移系统,为该菌的生物合成基因改造奠定了基础。Objective： The gene transfer system for Ascomycin producing strain Streptomyces hygroscopicus FIM260840 was established,which was used for gene disruption and expression of foreign genes. Method： Intergeneric genetic transfer system was constructed by conjugation with plasmid p SET152,and factors of influence for conjugation efficiency were investigated and optimized. Result： The conjugons could be screened by 12. 5 μg / m L Apramycin effectivity. PCR verification revealed that exogenous plasmid was successfully integrated in the chromosomal DNA of FIM260840. The resistant ability of conjugants to apramycin are more than 400μg / m L. The continuous passageculture experiment showed that transformed p SET152 of conjugants could be stably inherited. Conclusion： The efficient and simple gene transfer system for producing strain Streptomyces hygroscopicus FIM260840 was established,which laid the foundation for further biosynthetic gene modification with the strain.

关 键 词：子囊霉素 吸水链霉菌 P SET152 接合转移 遗传稳定性 

分 类 号：Q784[生物学—分子生物学]