解脂亚罗酵母原生质体的制备与再生条件
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摘要
使用纤维素酶和蜗牛酶复合酶液进行处理,并用β-巯基乙醇预处理,采用甘露醇作为渗透压稳定剂,考察菌龄、酶质量浓度、酶解温度和酶解时间对解脂亚罗酵母原生质体形成和再生的影响。实验得出结论:对于原生质体形成率影响因素来说,理论的最佳形成条件为:菌龄36 h,酶质量浓度为2.5mg/m L,酶解温度为25℃,酶解时间为8 h;对于原生质体再生率影响因素来说,理论的最佳再生条件为:菌龄48 h,酶质量浓度为2 mg/mL,酶解温度为25℃,酶解时间为8 h。而综合形成率和再生率共同影响因素顺序为:菌龄>酶质量浓度>酶解温度>酶解时间,最佳制备条件为:菌龄36 h,酶质量浓度2mg/mL,酶解温度25℃,酶解时间8 h。
The thallus of Yarrowia lipolytica was treated with cellulase and snail enzyme complex enzyme solution.It was pretreated with beta-mercaptoethanol and mannitol was used as the osmotic stabilizer.The effects of fungus age,enzyme concentration,enzymolysis temperature and enzymolysis time on protoplast formation and regeneration of Yarrowia lipolytic were investigated.It is concluded that the optimum conditions for protoplast formation were as follows:The fungus age was 36 h,the enzyme concentration was 2.5 mg/mL,the enzymolysis temperature was 25 ℃,and the enzymolysis time was 8 h.For the factors affecting protoplast regeneration,the best regeneration conditions were:The fungus age was 48 h,the enzyme concentration was 2 mg/mL,the enzymolysis temperature was 25 ℃,and the enzymolysis time was 8 h.And the order of the common factors affecting the formation rate and regeneration rate was:fungus age > enzyme concentration > enzymolysis temperature > enzymolysis time.The optimum preparation conditions were:the age of bacteria was 36 h,the enzyme concentration was 2 mg/mL,the enzymolysis temperature was 25 ℃,and the enzymolysis time was 8 h.
The thallus of Yarrowia lipolytica was treated with cellulase and snail enzyme complex enzyme solution.It was pretreated with beta-mercaptoethanol and mannitol was used as the osmotic stabilizer.The effects of fungus age,enzyme concentration,enzymolysis temperature and enzymolysis time on protoplast formation and regeneration of Yarrowia lipolytic were investigated.It is concluded that the optimum conditions for protoplast formation were as follows:The fungus age was 36 h,the enzyme concentration was 2.5 mg/mL,the enzymolysis temperature was 25 ℃,and the enzymolysis time was 8 h.For the factors affecting protoplast regeneration,the best regeneration conditions were:The fungus age was 48 h,the enzyme concentration was 2 mg/mL,the enzymolysis temperature was 25 ℃,and the enzymolysis time was 8 h.And the order of the common factors affecting the formation rate and regeneration rate was:fungus age > enzyme concentration > enzymolysis temperature > enzymolysis time.The optimum preparation conditions were:the age of bacteria was 36 h,the enzyme concentration was 2 mg/mL,the enzymolysis temperature was 25 ℃,and the enzymolysis time was 8 h.
引文
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[5]顾蕾,陆玲,袁生.红酵母原生质体制备及其紫外诱变育种的研究[J].食品工业科技研究与探讨,2004(4):60-65.
[6]CHENG Y L,BELANGER R R.Protoplast preparation and regeneration from spores of thebiocontrol fungus Pseudozyma flucculosa[J].FEMS Microbiology Letters,2000,190:287-291.
[7]喻欢.微生物絮凝剂及细胞融合技术处理焦化废水中酚氰的研究[D].南昌:南昌大学,2008.
[8]陈春涛,王岁楼.产类胡萝卜素酵母菌原生质体的制备再生与诱变[J].生物技术,2002(12):23-25.
[2]陈三凤,刘德虎.现代微生物遗传学[M].北京:化学工业出版社,2003:56-59.
[3]GEROLD B,CLAUDE G.Physiology and genetics of the dimorphic fungus Yarrowia lipolytica[J].Fems Microbiology Reviews,1997,19:219-237.
[4]BARNETT J A,PAYNE R W,YARROW D.Yeasts characteristics and identification[M].Cambridge:Cambridge University Press,1990:683-684.
[5]顾蕾,陆玲,袁生.红酵母原生质体制备及其紫外诱变育种的研究[J].食品工业科技研究与探讨,2004(4):60-65.
[6]CHENG Y L,BELANGER R R.Protoplast preparation and regeneration from spores of thebiocontrol fungus Pseudozyma flucculosa[J].FEMS Microbiology Letters,2000,190:287-291.
[7]喻欢.微生物絮凝剂及细胞融合技术处理焦化废水中酚氰的研究[D].南昌:南昌大学,2008.
[8]陈春涛,王岁楼.产类胡萝卜素酵母菌原生质体的制备再生与诱变[J].生物技术,2002(12):23-25.