绿色木霉原生质体制备与再生条件的探索

研究菌丝体培养时间、酶组合、酶浓度、酶解时间、酶解温度、稳渗剂类型、再生培养方式等对绿色木霉原生质体制备及再生的影响。结果表明,绿色木霉原生质体制备的优化条件为：菌丝体培养60h,用1.5%溶菌酶＋0.5%蜗牛酶的混合酶液,在pH值6.0～6.5,30～35℃,酶解4h,以0.7mol/L甘露醇为稳渗剂,原生质体的产量可达1.68×107个/ml;原生质体再生的优化条件为：PDA为再生培养基,以0.5mol/L蔗糖为稳渗剂进行双层固体培养,原生质体的再生率为6.05%～6.12%。

Exploration on Preparation and Regeneration Conditions for Protoplast of Trichoderma viride

The effects of culture time of mycelium,enzyme combinations,enzyme concn.,enzymolysis time,hydrolysis temperature,osmotic stabilizer types,regeneration methods etc on preparation and regeneration of protoplast of Trichoderma viride were studied.The results showed that the optimum condition for preparation of protoplast of T.viride was culture time of mycelium of 60 h,enzyme combination of 1.5% lysozyme +0.5% snail enzyme,hydrolysis temperature of 30-35 ℃ and enzymolysis time of 4 h with pH value of 6.0-6.5 and osmotic stabilizer of 0.7 mol/L mannitol.Under this condition the yield of protoplast could reach 1.68×10~7/ml.The optimum condition for regeneration of protoplast of T.viride was PDA as the regeneration medium with osmotic stabilizer of 0.5 mol/L sucrose and regeneration method of double-layer solid culture.Under this condition the regeneration rate of the protoplast was 6.05%-6.12%.