重离子诱变选育高产核黄素枯草芽孢杆菌

Screening of high-yield riboflavin Bacillus subtilis strain by heavy-ion beam irradiation

研究以产核黄素的枯草芽孢杆菌C0为出发菌株,经^(12)C^(6+)重离子辐射诱变后,以8-氮鸟嘌呤作为拮抗物进行高产菌株的筛选,初步获得60个诱变菌株,进一步筛选得到4株高产菌株,最后经摇瓶发酵,发现4株诱变菌株中B13的核黄素产量最高为(401.8±9.2)mg/L,比原始菌株提高18.5%。经二次重离子诱变选育后,获得一株高产菌株BII7,核黄素产量达到(818.8±17.3)mg/L,比一次诱变菌株B13提高77.7%,表明重离子辐射对产核黄素的枯草芽孢杆菌具有良好的诱变效果。对诱变菌株BII7进行培养温度、pH值及转速等条件的优化,最适培养条件为37℃、pH 7.7、220 r/min。在5 L双联发酵罐中扩大培养高产菌株BII7,37℃、pH 7.7及溶氧浓度40%时,发酵时间缩短12 h,核黄素产量达1385.3 mg/L,比摇瓶培养提高69.2%,该诱变菌株具有较好的放大生产能力。6次传代培养实验表明该诱变高产菌株具有稳定的遗传特性。

The parent riboflavin-producing strain Bacillus subtilis C0 was treated with heavy-ion beam irradiation,then it was screened the mutagenic strains with 8-azaguanine.In further study,4 high-yield strains were initially obtained from 60 mutant strains.After shake flask fermentation,the highest yield of mutant strain B13 reached(401.8±9.2)mg/L,which was 18.5%higher than that of the original strain.The new mutant strain BII7 was screened after second time of heavy-ion beam irradiation,and the riboflavin yield achieved(818.8±17.3)mg/L which was 77.7%higher than that of the mutation strain B13.The heavy-ion beam irradiation had good mutagenic effect.The optimal cultivation conditions of strain BII7 were:initial pH 7.7,culture temperature 37℃,and rotation speed 220 r/min.The riboflavin yield of mutant strain BII7 was 1385.3 mg/L in 5 L fermenter(37℃,pH 7.7 and dissolved oxygen concentration 40%),which was 69.2%higher than that in shake flask.The fermentation time was shortened by 12 h.The results showed that the strain BII7 had good fermentation ability and stable genetic characteristics.