【摘要】 目的利用原生质体诱变方法从一株链霉菌原始菌株Streptomyces sp. H-323(CGMCC 4827)选育阿霉素高产突变菌株,以提高阿霉素的发酵单位,并对发酵产品的质量进行了初步研究。方法通过研究不同甘氨酸浓度、溶菌酶酶解时间、酶解温度、酶浓度对H-323原生质体形成的影响,确定了原生质体的最佳制备和再生条件。通过阿霉素抗性选育,诱变原生质体筛选阿霉素高产突变菌株。最后通过高效液相、核磁等波谱分析法初步确定发酵法生产的阿霉素产品的质量。结果制备Streptomyces sp. H-323原生质体的最佳条件:种子培养基中甘氨酸浓度0.5%、溶菌酶浓度3mg/mL、酶解时间60min、酶解温度30℃。通过原生质体诱变结合阿霉素抗性选育,得到一株高产突变株,阿霉素发酵单位由出发菌株的300mg/L提高到700mg/L。通过波谱分析,发酵产物达到国外注册标准。结论通过原生质体诱变和抗性选育大大提高了阿霉素的产量,为高产阿霉素产业化提供方法支持。更多还原

【Abstract】 Objective To increase the production of doxorubicin,a high doxorubicin-producing strain was screened through protoplast mutation from original strain Streptomyces sp.H-323(CGMCC 4827).The quality of the fermentation product was studied.Methods The optimum conditions,including concentration of glycine in seed medium,concentration of lysozyme,enzymolysis time and enzymolysis temperature,which affected the formation and regeneration of the protoplasts were studied.Doxorubicin-resistance selection and protoplast induction were performed to breed a high doxorubicin-producing mutant strain.Spectral analysis of HPLC and NMR were performed to determine the quality of the fermentation product.Results The optimum conditions of preparing protoplast were as follows: seed medium containing 0.5% glycine,the mycelia were processed with 3mg/mL lysozyme at 30°C for 60min.A high doxorubicin-producing mutant strain was selected by protoplast mutation with doxorubicin-resistance method.The doxorubicin production of the mutant strain was remarkably increased to 700mg/L compared to the production 300 mg/L of original strain.The quality of the fermentation product meets the clients’ standard according to the spectral analysis.Conclusion Protoplast mutation and doxorubicin-resistance selection is a reasonable method to breed high doxorubicin-producing strain.This study provides an useful method support for industrialization of doxorubicin by fermentation.更多还原