通过获得链霉素抗性基因(str)突变株筛选梅岭霉素高产菌株

Acquisition of streptomycin-resistant (str) mutants screens high-yield meilingmycin strain in Streptomyces nanchangensis

本文通过链霉素对梅岭霉素 (meilingmycin)产生菌南昌链霉菌NS 41 80菌株孢子致死浓度的测定 ,采用诱变剂EMS四种不同诱变剂量对菌株的孢子进行诱变处理 ,诱变处理的孢子涂布在含链霉素致死浓度的高氏平板上 ,获得了大量的链霉素抗性基因 (str)突变株。然后从链霉素抗性基因突变株进一步筛选到梅岭霉素高产菌株 80 5 .11 2 2 1,在摇瓶条件下 ,只产梅岭霉素不产南昌霉素 ,梅岭霉素活性效价达 15 0 0 μg/ml,比出发菌株NS 41 80的摇瓶发酵效价 85 5 μg/ml提高了 77.9%,该菌株连续传代六代进行摇瓶发酵 ,其F2 代和F3 代梅岭霉素发酵效价稳定 ,F4代至F6代随着传代数增加 ,其梅岭霉素发酵效价急速下降。通过EMS诱变剂量分别与抗药性突变率和链霉素抗性基因突变株产梅岭霉素产量的产势统计分析表明 ,菌株抗药性突变与产抗生素突变密切相关 ,产抗生素突变的EMS诱变剂量高于链霉素抗性基因突变诱变剂量。在 0 .0 3mol/L的EMS剂量作用下 ,菌株致死率为 99.43 %,而抗药性突变率为 0 .0 44 0 %,建立了梅岭霉素产生菌链霉素抗性基因突变筛选方法 ,为南昌链霉菌高产菌种选育研究作了有益的尝试 。

After testing the resistance of streptomycin to the strain NS 41 80 which is the meilingmycin producer-- Streptomyces nanchangensis , a mumber of streptomycin resistant ( str ) mutants were screened after the spores regenerated on the lethal media when they were treated with 4 different concentrations of super mutangent EMS. We obtained the high yield strain 80 5.11 221 from these str mutants which only produced meilingmycin and no nanchangmycin in the rotation flask experiments. The potency of 80 5.11 221 was 1521μg/ml which was 77.9% higher than that of 855μg/ml. After six generations of shake flask fermentation experiment of 80 5.11 221, the potencies of F 2 and F 3 is stable but F 4, F 5 and F 6 decreases hastily. By statistic analysis of different mutation approaches, the lethal frequency rate of spores using EMS as mutagen at a dosage of 0.03mol/L was 99.43%, while the streptomycin resistant mutation rate was 0.0440% . The rational screening approach using antibiotic to select high producing meilingmycin strain was established.