盐霉素高产菌株的筛选研究  

The study of screening for high-salinomycin-yield mutants

作　　者：胡莉莉[1] 梁剑光[1,2] 陈中兵 郭美锦[1] 储炬[1] 庄英萍[1] 张嗣良[1] 

机构地区：[1]华东理工大学生物反应器工程国家重点实验室,上海200237 [2]浙江升华拜克生物技术股份有限公司,湖州313220

出　　处：《中国抗生素杂志》2015年第12期906-912,共7页Chinese Journal of Antibiotics

基　　金：国家重大科学仪器设备开发专项(No.2012YQ150087);江苏省高校自然科学基金项目(No.11KJB180001)

摘　　要：目的优化高通量筛选盐霉素高产菌株流程,高通量筛选出具有遗传稳定性的产盐霉素高产菌株。方法以核糖体工程理论为基础,常压室温等离子体(atmospheric and room temperature plasma,ARTP)及高通量筛选技术(high-throughput screening,HTS)对产盐霉素的白色链霉菌(Streptomyces albus)S12诱变筛选,并以琼脂块扩散法为基础建立一种快速检测盐霉素的方法应用于高通量筛选过程初筛。结果获得具有氯霉素和四环素双重抗性的高产菌株Tet30Chl25。摇瓶发酵实验结果表明:突变株Tet30Chl25盐霉素效价由初始菌株的16306U/m L提高到34712U/m L,提高了112%,且遗传稳定性良好。同时建立一种快速检测盐霉素的方法——琼脂块扩散法并应用于高通量筛选过程初筛中,缩短筛选周期12d,优化了筛选流程。结论首次综合运用核糖体工程推理选育技术、常压室温等离子体(atmospheric and room temperature plasma,ARTP)及高通量筛选技术(highthroughput screening,HTS)对产盐霉素的白色链霉菌(S.albus)诱变筛选,是很好的菌种选育策略,适合工业菌种的选育。Objective To optimize the process of high-yield strains producing salinomycin by high-throughput screening, obtain high-yield and genetic stable mutants of Streptomyces albus and improve the titer of salinomycin. Methods The ribosome engineering, the atmospheric and room temperature plasma （ARTP） and high-throughput screening （HTS） were firstly comprehensively applied in the screening of high-yield strains of salinomycin, and a fast test method has been established to detect high salinomycin titer of mutants based on the agar block method. Results A mutant Tet30Ch125 with tetracycline and chloramphenicol resistance was selected by high-throughput screening. In the shaking flask culture, the mutant Tet30Ch125 was hereditarily stabilized and its yield of salinomycin increased to 34,712U/mL compared to 16,306U/mL produced by the original strain, namely, increased by 112%. The subculture experiments indicated that the hereditary characteristics of the mutant Tet3~ChF5 was relatively stable. In addition, the agar block method shortened the screening period by 12 days so as to save the mutant breeding duration when high- throughput screening was used in the procedure of high-yield strains of salinomycin. Conclusion The ribosome engineering, the atmospheric and room temperature plasma （ARTP） and high-throughput screening（HTS） were firstly integrated in the screening of high-yield strains of salinomycin, which was an effective strategy for strain breeding. Therefore, this combined method can be applied in the mutation and breeding of industrial strains.

关 键 词：琼脂块扩散法 核糖体工程 ARTP诱变技术 高通量筛选技术 盐霉素 

分 类 号：R978.1[医药卫生—药品]