小白链霉菌(Streptomyces albulus)的原生质体制备研究

Study on the protoplasts formation of Streptomyces albulus

广谱天然防腐剂ε-多聚赖氨酸产生菌Streptomyces albulus的育种工作目前仅处于常规育种水平,为对其进行原生质体技术的育种改造,对影响S.albulus原生质体制备的几个重要因子进行了详细研究。测定了S.albulus的生长曲线,确定制备原生质体的最佳菌龄为稳定期初期即培养40h;S发酵液中添加2·0%甘氨酸可使原生质体形成率最高;以破坏链霉菌细胞壁效果最好的溶菌酶进行酶解,确定最佳酶解温度为为35℃、最佳酶解时间为180min、最佳溶菌酶浓度为2.0mg·mL-1;在以上最佳原生质体制备条件下可获得纯净且量多的S.albulus原生质体,形成量达1.74×108cfu·mL-1,为S.albulus原生质体的进一步遗传育种奠定了良好的基础。

S. albulus that produces the broad spectrum peptide biopreservative is just at conventional breeding level now. In order to breed using protoplasts, several factors affecting the protoplasts formation of S. albulus were studied. The growth curve was determined. The optimum cell -age for protoplasts formation was in the early stationary phase, namely 40 h. The maximum yield of protoplasts were obtained, when pretreated by 2.0% glycin in the S culture medium. The optimum enzyme condition is digesting at 35℃ for 180 min with 2.0 mg·mL^-1 lysozyme that is the best enzyme to digest cell wall of streptomycete. On the optimum conditions, numerous and pure protoplasts were obtained. The highest yield of protoplasts amounted to1.74×10^8cfu·mL^-1 . The foundation for genetic breeding of S. albulus has been laid by using the protoplasts farther.