外源vpl1基因对香菇双核菌丝体木质素降解酶活性和基因表达的影响  

Effects of exogenous vpl1 gene on lignin-degrading enzyme activities and gene expression in the dikaryotic mycelia of Lentinula edodes

作　　者：游潇轩 宋晓霞[2] 戈永杰 刘建雨 张美彦[2] 尚晓冬[2] 宋春艳[2] YOU Xiaoxuan;SONG Xiaoxia;GE Yongjie;LIU Jianyu;ZHANG Meiyan;SHANG Xiaodong;SONG Chunyan(College of Food Science&Technology,Shanghai Ocean University,Shanghai 201306,China;National Engineering Research Center of Edible Fungi,Key Laboratory of Edible Fungal Resources and Utilization(South),Ministry of Agriculture and Rural Affairs of the People’s Republic of China,Institute of Edible Fungi,Shanghai Academy of Agricultural Sciences,Shanghai 201403,China)

机构地区：[1]上海海洋大学食品学院,上海201306 [2]上海市农业科学院食用菌研究所农业农村部应用真菌资源与利用重点实验室国家食用菌工程技术研究中心,上海201403

出　　处：《菌物学报》2024年第7期66-78,共13页Mycosystema

基　　金：上海市食用菌产业技术体系[沪农科产字(2023)第9号]。

摘　　要：为探索刺茎侧耳多功能过氧化物酶vpl1基因在香菇菌丝体中的异源表达及功能,对比分析了野生型双核菌丝体与拥有1个或2个转化子的双核菌丝体在木质素降解酶活性和基因表达上的差异。首先,经过转化子筛选和功能验证,筛选出3个具有vpl1基因的香菇t-Asm136单核转化子。其次,将3个单核转化子与其野生型w-Asm136单核菌丝体分别与实验室已有的香菇t-Bsm83单核转化子与其野生型w-Bsm83单核菌丝体进行单单杂交,获得1个纯野生型双核菌丝体、4个单转化子双核菌丝体和3个双转化子双核菌丝体。最后,仅野生型、1个单转化子和2个双转化子双核菌丝体能在木屑培养基上正常生长。这4个双核菌丝体在木屑培养基上Lac、MnP和VP酶活和对应基因的表达量不同,其中vpl1的导入使双核转化子vpl1基因上调了2-12倍,MnP酶活上调13-30倍,mnp2a表达量上调4-18倍,mnp2b表达量上调6-11倍。证实了vpl1的导入能够增强MnP的酶活力和基因的表达水平。To explore the heterologous expression and function of Pleurotus eryngii versatile peroxidase vpl1 gene in the mycelium of Lentinula edodes,the differences of lignin-degrading enzyme activity and gene expression between the dikaryotic mycelia without vpl1 transformant and those containing one or two vpl1 transformants were compared.Three mononuclear transformants(L.edodes t-Asm136)containing vpl1 gene were obtained through genetic transformation screening and gene function verification.The mycelia of the three mononuclear transformants(t-Asm136)and their wild-type(w-Asm136)were respectively mon-mon crossed with the mycelia of mononuclear transformant t-Bsm83 and its wild-type w-Bsm83 containing and without vpl1 previously obtained by our laboratory.One dikaryotic wild-type strain without transformant,four dikaryotic strains containing one transformant,and three dikaryotic strains containing two transformants were obtained.It was found that’s only the dikaryotic wild-type strain without transformant,one dikaryotic strain containing one transformant,and two dikaryotic strains containing two transformants could grow normally in sawdust substrate.The enzyme activities of Lac,MnP and VP and the expression levels of related genes were different among these four dikaryotic strains.The introduction of vpl1 can up-regulate the vpl1 gene by 2-12 times,the MnP enzyme activity by 13-30 times,the mnp2a expression by 4-18 times,and the mnp2b expression by 6-11 times,confirming that the introduction of vpl1 can enhance the enzyme activity and gene expression of MnP.

关 键 词：香菇 遗传转化 多功能过氧化物酶 异源表达 

分 类 号：S646.12[农业科学—蔬菜学]