【摘要】 采用 0 .6%纤维素酶加 0 .6%蜗牛酶的混合酶由扩展青霉 K4 0中获得大量的原生质体 ,并比较了菌龄、二硫苏糖醇 ( DTT)预处理方式、酶解时间、温度、 p H、培养基成分及稳定剂等因素对原生质体的形成和再生的作用 .选出制备原生质体的最适条件 :0 .6%纤维素酶 +0 .6%蜗牛酶 ,在 0 .6mol/L Na Cl+0 .3 %Ca Cl2 ,DTT预处理 0 .5h,菌龄为 1 9～ 2 0 h,p H 5.4 ,2 8℃酶解 3～ 3 .5h,原生质体产量可达 2 .3 6× 1 0 7个/ml,实现再生 ,再生率达 70 %～ 80 % .并对原生质体的释放和再生方式进行跟踪观察更多还原

【Abstract】 A fast and reliable method of obtaining protoplasts from Penicillium expansum K40 mycelia is report here.A protoplast yield of2 .3 6× 1 0 7/ml can be obtained in3～ 3 .5 hours under observed optimal conditions which are described as follows:Using 1 9～ 2 0 hours old wheatbran agar medium cultures of Penicillum expansum K40 ,0 .6% cellulase plus 0 .6% snailase,p H 5 .4without any buffer,petri dishes of6cm diameter as incubating container with3 ml of mycelia and enzyme mixture;mycelia pretreated by0 .0 1 mol/L DTT for3 0 minutes,0 .6mol/L Na Cl plus0 .3 % Ca Cl2 as osmotic stabilizer;temperature range 2 5℃～ 3 0℃ . Several forms of protoplasts release and regeneration were observed. The conditions for protoplast regeneration were also investigated. The protoplastwere inoculated on2 % agarplates overlaid with0 .8% agar,afterincubated at2 5℃～ 3 0℃ for40 hours,regeneration occurred and colonies were formed.The rate of regeneration over70 % .更多还原