松茸原生质体制备、再生条件优化及释放方式观察  

Optimizing conditions of protoplast preparation and regeneration of Tricholoma matsutake and its release modes

作　　者：张丽[1,2] 郭成金[1,2] 

机构地区：[1]天津师范大学生命科学学院 [2]天津市动植物抗性重点试验室,天津300387

出　　处：《浙江农业学报》2014年第4期941-947,共7页Acta Agriculturae Zhejiangensis

基　　金：国家星火计划项目(2012GA610002)

摘　　要：借助正交设计法对松茸原生质体制备与再生进行研究,结果表明,松茸原生质体释放方式主要有4种,分别为顶端释放、侧位释放、菌丝段端位释放及原位释放。原生质体最佳制备体系是:菌龄2 d,以0.6 mol·L-1的KCl为渗透压稳定剂,10 L+10 S+10 C的酶系统34℃酶解2 h,最终制备率达到2.75×107个·mL-1;最佳再生体系为:菌龄2 d,以MgSO4·7H2O为渗稳剂,10 L+10 S的酶液30℃酶解4 h,在以蔗糖为渗稳剂的再生培养基上再生率最高,可达41.6%。This study aimed to explore the protoplast preparation and regeneration conditions of Tricholoma matsutake by orthogonal experiment .The results showed that there were four release modes of T.matsutake protoplasts：release from top,release from the side,hyphal fragment end release and release in situ .The optimal protoplast preparation condition was selecting mycelium cultured in liquid medium for 2 d, for enzymolysis in mixed enzyme solution containing 1% cellulose , 1% snailase and 1% lywallzyme at 34℃ for 2 h, with 0.6 mol· L^-1 KCl as osmotic stabilizer,and the obtained preparation rate reached 2.75 ×107protoplasts·mL^-1.The optimal regeneration condition was using mycelium cultured in liquid medium for 2 d,for enzymolysis in enzyme solution containing 1%lywallzyme and 1% snailase at 30℃ for 4 h,with 0.6 mol· L^-1 MgSO4·7H2O as osmotic stabilizer,and the regeneration rate reached 41.6%.

关 键 词：松茸 原生质体 释放方式 正交设计 再生率 

分 类 号：Q813.2[生物学—生物工程]