Objective To determine the optimal conditions of protoplast preparation and regeneration of Brevibacterium flavum which produces glutamine highly.Methods we used lysozyme to hydrolyze Brevibacterium flavum which had been pretreated with penicillin and glycine,then researched the effects of cell age,concentration and pretreatment time of penicillin and glycine,enzyme concentration,enzymolysis time,enzymolysis temperature on protoplast preparation and regeneration.Results The optimal conditions of protoplast preparation and regeneration were as follows:pretreating Brevibacteriumflavum at the early time of exponential phase,the optimum concentration of penicillin of 0.4 u/mL,2 ％ glycine,preconditioning time of 2 h,enzyme concentration of 1 mg/mL,enzymolysis time of 12 h,enzymolysis temperature of 37 ℃.Conclusion Under the optimal conditions,the protoplast formation rate can be 99.71％,the regeneration rate can be 16.52 ％.%目的 研究高产谷氨酰胺的黄色短杆菌原生质体制备和再生的最佳条件.方法 用青霉素和甘氨酸预处理黄色短杆菌后,用溶菌酶酶解,研究菌龄、青霉素和甘氨酸预处理浓度及时间、酶浓度、酶解时间、酶解温度等条件对原生质体制备和再生的影响.结果 原生质体制备和再生的最佳条件为:预处理处于对数生长早期的黄色短杆菌,青霉素最适浓度0.4 u/mL,甘氨酸浓度2％,预处理时间2h,酶浓度1 mg/mL,酶解时间12h,酶解温度37℃.结论 在最佳条件下原生质体形成率达99.71％,再生率达16.52％.
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