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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 study the optimum conditions for protoplast preparation and regeneration of Streptomyces nosinophilus. Methods Lysozyme was used to remove the cell wall to prepare protoplasts. Various influencing factors of protoplast preparation and regeneration were investigated. Results The conditions of protoplast preparation were determined: seed culture medium was used in primary culture for 30 h and the volume fraction of transplanting was 5%; secondary culture was carried out with R2YE medium for 32 h, the optimum glycine concentration 6.0 gL-1, the optimal concentration of lysozyme was 1.5 gL-1, the enzymolysis time was 60 min and the regeneration rate of protoplasts reached 5.3%. Conclusion The above conditions are the most suitable conditions for the preparation and regeneration of Streptomyces protoplast. The establishment of this condition laid a foundation for the mutation breeding of Streptomyces protoplast.