马爱民 ; 贺冬梅 ; 潘迎捷
上海市农业科学院食用菌研究所 ; 农业部食用菌遗传育种重点实验室 上海 201106
【中文摘要】 通过对溶壁酶组成及浓度,渗透压稳定剂、菌丝培养方法等研究,建立了有效的双孢蘑菇原生质体制备与再生系统。采用1.5％Lywallzyme、0.6M KCl为渗透压稳定剂,25℃酶解4小时,原生质体的产量可达到107个/ml;纯化后的原生质体在0.6M蔗糖为渗透压稳定剂的PDMA培养基中,25℃恒温培养5～7天,再生率达1%以上,根据原生质体再生菌落出现时间的先后顺序,并以菌落形态特征、菌丝生长速度。羧甲基纤维素酶(Cx酶)活性及子实体形成能力上的差异为鉴定同核体的主要标记,从两种形态类型,不同来源的12个双孢蘑菇菌株中均分离到同核原生质体,同核率为2％～15%,平均为11.75％。
【英文摘要】 Based on testing of different combinations and concentration of lysozymes, osmotic stabilizers and different methods of mycelium incubation, an efficient system of protoplast preparation and regeneration in Agaricus bisporus was established in present paper. It was proved that after the mycelium being digested with 1.5% lywallzyme in 0.6M KCl for 4h at 25℃, the protoplast yield reached 107/ml, and that after the purified protoplasts being incubated in PDMA medium with 0.6M sucrose as osmotic stabilizer for 5-7 days at 25℃, the regeneration rate was over 1%. Based on different regeneration time, using the difference of colonial morphology, activity of Cx enzyme and fruiting ability as main markers for identification of homokaryotic protoplast, the homokaryotic protoplasts were isolated from protoplasts of 12 strains with a rate of 2% to 15%, averaging out at 11.75%.
【中文关键词】 双孢蘑菇; 同核原生质体; 分离与鉴定
【英文关键词】 Agaricus bisporus; Homokaryotic protoplast; Isolation and identification
【基金】上海市科委资助项目
【文献出处】 食用菌学报,Acta Edulis Fungi,编辑部邮箱,1995年03期 【DOI】CNKI:SUN:SYJB.0.1995-03-000