诺西肽产生菌活跃链霉菌的原生质体制备与再生

Protoplasts preparation and regeneration of Streptomyces actuosus:the producer of nosiheptide

目的研究诺西肽产生菌活跃链霉菌的原生质体制备与再生的最适条件。方法使用溶菌酶脱去细胞壁制备原生质体,并考察原生质体制备和再生的各种影响因素。结果确定了原生质体制备的条件:一级培养采用种子培养基,培养30 h,转种量的体积分数为5%;二级培养采用R2YE培养基,培养时间为32 h,最适甘氨酸质量浓度为6.0 g.L-1,最佳溶菌酶质量浓度为1.5 g.L-1,酶解时间为60 min,原生质体再生率达到5.3%。结论上述条件为活跃链霉菌原生质体制备与再生的最适条件,该条件的建立为活跃链霉菌原生质体的诱变育种奠定了基础。

Objective To obtain the optimal conditions for protoplasts formation and regeneration of Streptomyces actuosus：the producer of nosiheptide.Methods Protoplasts of Streptomyces actuosus were prepared by lysozyme digestion;the conditions for protoplasts formation and regeneration were optimized through the investigation on the factors influencing the preparation and regeneration of protoplasts.Results The conditions for the preparation of protoplasts were confirmed as following：primary mycelia were cultivated for 30 h in seed medium,the inoculation volume of seed culture to second phase culture was 5%;then the seed culture was transferred to the R2YE liquid medium with 6.0 g·L-1 glycine for the secondary mycelia cultivation.After 32 h cultivation,the mycelia were collected,washed and lysed by lysozyme at the optimal concentration of 1.5 g·L-1 for 60 min.The obtained protoplast regeneration rate was 5.3%.Conclusions Above conditions are the optimum conditions for the protoplasts formation and regeneration of Streptomyces actuosus.The establishment of these optimum conditions lay a foundation for the mutational breeding of Streptomyces actuosus.