灰葡萄孢菌BcGLS基因的克隆、生物信息学分析及逆境胁迫下转录表达分析  

Cloning and bioinformatic analysis of BcGLS gene in Botrytis cinerea and transcriptional expression analysis

作　　者：孟利娟 昝新艺[1] 杨雨蒙 张子盈 崔凤杰[1] 田贞乐[2] MENG Lijuan;ZAN Xinyi;YANG Yumeng;ZHANG Ziying;CUI Fengjie;TIAN Zhenle(School of Food and Biological Engineering,Jiangsu University,Zhenjiang 212013,China;Jiangsu Alphay Biotechnology Co.Ltd.,Nantong 226009,China)

机构地区：[1]江苏大学食品与生物工程学院,江苏镇江212013 [2]江苏安惠生物科技有限公司,江苏南通226009

出　　处：《食品与发酵工业》2024年第18期169-176,I0001,I0002,共10页Food and Fermentation Industries

基　　金：江苏省高等学校基础科学(自然科学)研究面上项目(23KJB210005);江苏大学高级人才科研启动项目(5501360015)。

摘　　要：β-1,3-葡聚糖合成酶(β-1,3-glucan synthase,GLS)催化合成的细胞壁核心组分β-1,3-葡聚糖对真菌的生存和毒力至关重要,该酶是研发抗菌药物的重要作用靶点。为了了解β-1,3-葡聚糖合成酶基因(BcGLS)在灰葡萄孢菌生存和侵染进程中的功能,该研究通过克隆获得BcGLS全长序列,序列全长7137 bp,编码1932个氨基酸,生物信息学分析结果显示,其编码蛋白(BcGLSp)理论相对分子质量为220.05 kDa,理论等电点为7.97;亚细胞预测其定位于细胞膜,存在17个跨膜结构,不存在信号肽,为非分泌蛋白;含有176个磷酸化位点,为亲水性蛋白。保守结构域分析结果表明,BcGLSp含有一个β-1,3-葡聚糖合成酶结构域和一个β-1,3-葡聚糖合成酶亚基FKS1,属于葡聚糖合成酶超家族。系统进化分析表明,灰葡萄孢菌BcGLS与植物致病菌如山茶叶杯菌、景天白粉菌等的葡聚糖合成酶亲缘关系近。不同逆境胁迫条件(氯化钠、十二烷基硫酸钠、过氧化氢)显著抑制灰葡萄孢菌菌丝生长,实时定量PCR进一步分析显示BcGLS在不同非生物胁迫条件下均上调表达,表明BcGLS基因的转录表达与环境压力密切相关。该研究为深入探讨BcGLS在灰葡萄孢菌生存和侵染进程中的基因功能和灰葡萄孢菌的新型防治策略提供理论材料和实践依据。β-1,3-glucan synthase(GLS)catalyzes the synthesis of cell wall core componentβ-1,3-glucan which is essential for fungal survival and virulence,and is the important target for antifungal agent development.To explore its gene function in the survival and infection process of Botrytis cinerea,this study primarily cloned the full-length sequence ofβ-1,3-glucan synthase coding gene(BcGLS).Its total length was 7137 bp by encoding 1932 amino acids.Bioinformatic analysis indicated that BcGLS protein(BcGLSp)had a theoretical relative molecular weight of 220.05 kDa and a theoretical isoelectric point of 7.97.Subcellular prediction of BcGLSp was located in the cell membrane with 17 transmembrane regions,it had no signal peptide and was a non-secreted protein.BcGLSp was a hydrophilic protein with 176 phosphorylation sites.The results of conservative domain analysis revealed that BcGLSp contains oneβ-1,3-glucan synthase component and oneβ-1,3-glucan synthase FKS1 domain,belonging to glucan-synthase superfamily.Phylogenetic analysis displayed that BcGLS had close genetic relationship with those from plant pathogens such as Ciborinia camelliae and Erysiphe neolycopersici.Different abiotic stresses(NaCl、SDS、H_(2)O_(2))significantly reduced Botrytis mycelial growth rate,and real-time PCR further showed that the expression of BcGLS was up-regulated under these stresses,indicating that the transcriptional expression of BcGLS was closely associated to environmental stresses.This study provides theoretical materials and practical basis for further research on the function of BcGLS gene in the survival and infection process of B.cinerea and the novel control strategies of B.cinerea.

关 键 词：灰葡萄孢菌 BcGLS 生物信息学 逆境胁迫 转录表达 

分 类 号：S432.44[农业科学—植物病理学]