中文摘要：

目的建立新磷氮霉素A产生菌金黄色链霉菌AGR0001（Streptomycesauratus AGR0001）的遗传操作体系，为阐明其生物合成的调控和后修饰机制，并通过遗传改造提高其产量和获得更多的新化合物奠定基础，同时可为其它链霉菌的遗传操作研究提供新的参考。方法筛选了SauratusAGR0001生长、产孢的最佳培养基，检测了其对11种不同抗生素的敏感性，探索了将外源DNA导入该菌的最适方法。结果ISP4和MS培养基分别是S. auratus AGR0001的最佳产孢和生长培养基；该菌对壮观霉素等7种抗生素敏感，对氨苄和羧苄西林具有抗性，对萘啶酮酸和阿伯拉霉素在低浓度有抗性，而在高浓度敏感；使用PEG介导的原生质体转化、电转菌丝体、接合转移等方法均能将外源DNA导入sauratusAGR0001中。利用接合转移将携带透明颤菌血红蛋白基因vhb（Vitreoscillahemoglobin）的自杀型质粒pSETl52导入S. auratus AGR0001中并整合至其染色体上进行表达，提高了发酵液的抗真菌活性。结论建立了S. auratus AGR0001的遗传操作体系。

英文摘要：

Objective To develop a genetic manipulation system in the neophoslactomycin A producer Streptomyces auratus AGR001 for further elucidating the regulatory and post modification mechanisms of phoslactomycin biosynthesis, which would facilitate further improving the production of phoslactomycin or obtaining other novel compounds through genetic engineering. The study will also provide clues for genetic manipulation in other streptomyces. Methods Different media for culturing S. auratus AGR0001 were tested to select the optimal medium for growth or generating spores. Resistance to 11 antimicrobial agents was measured. The optimal methods for introducing foreign DNA into the strain were also investigated. Results The optimal media for S. auratus AGR0001 spore generating and mycelium growth was ISP4 and MS, respectively. S. auratus AGR0001 was sensitive to spectinomycin, chloramphenicol, tetracycline, erythromycin, thiostreptone, rifampicin, kanamycin, and resistant to ampcilin and carbenicillin. For ampramycin and nalidixic acid, it was resistant at low concentrations （≤25μg/mL） and sensitive at higher concentrations （≥50μg/mL）. Foreign DNA could be introduced into this strain by using conjugation, mycelium electroporation, or PEG mediated protoplast transformation. The vhb gene from Vitreoscilla,cloned in the suicide plasmid pSET152, was introduced into S. auratus AGR0001 by using conjugation. The integration of vhb gene on the chromosome of S. auratus AGR0001 was confirmed. The fermentation products from the engineered strain exhibited improved antifungal activity. Conclusion The genetic manipulation system of S. auratus AGR0001 has been established.