刺葡萄STS基因克隆及其在不同光质下的表达水平  

Cloning of STS genes in Vitis davidii and their expression pattern under different light qualities

作　　者：许恒 赖恭梯 贺丽媛 李思雨 林俊璇 郭奥琳 赖谱富[2] 车建美[3] 陈桂信[1] 赖呈纯[2] XU Heng;LAI Gongti;HE Liyuan;LI Siyu;LIN Junxuan;GUO Aolin;LAI Pufu;CHE Jianmei;CHEN Guixin;LAI Chengchun(College of Horticulture,Fujian Agriculture and Forestry University,Fuzhou,Fujian 350002,China;Institute of Food Science and Technology,Fujian Academy of Agricultural Sciences/Key Laboratory of Processing of Subtropical Characteristic Fruits,Vegetables and Edible Fungi,Ministry of Agriculture and Rural Affairs of China,Fuzhou,Fujian 350003,China;Institute of Resources,Environment and Soil Fertilizer,Fujian Academy of Agricultural Sciences,Fuzhou,Fujian 350003,China)

机构地区：[1]福建农林大学园艺学院,福建福州350002 [2]福建省农业科学院农产品加工研究所/农业农村部亚热带特色果蔬菌加工重点实验室,福建福州350003 [3]福建省农业科学院资源环境与土壤肥料研究所,福建福州350003

出　　处：《福建农林大学学报（自然科学版）》2024年第4期465-473,共9页Journal of Fujian Agriculture and Forestry University:Natural Science Edition

基　　金：福建省自然科学基金项目(2021J05091);福建省公益类科研院所专项(2021R1032009);福建省财政专项——科技创新团队项目(CXTD2021018-2);福建省人民政府-中国农业科学院农业高质量发展超越“5511”协同创新工程项目(XTCXGC2021014)。

摘　　要：【目的】研究刺葡萄芪合酶(STS)基因的序列特征及其在不同光质处理下的表达水平,旨在为进一步解析STS基因通过响应光信号调控刺葡萄愈伤组织中白藜芦醇合成的分子机制提供依据。【方法】以刺葡萄愈伤组织为材料,利用逆转录PCR(RT-PCR)技术成功克隆获得了4个STS基因,对这些基因及其编码蛋白进行了生物信息学分析,并利用实时荧光定量PCR(RT-qPCR)检测其在不同光质下和不同培养阶段的表达水平。【结果】成功地从刺葡萄愈伤组织中克隆获得了VdSTS5、VdSTS6、VdSTS20、VdSTS21共4个基因,均含1179 bp完整开放阅读框(ORF),编码392个氨基酸残基,预测其编码的是亲水性、无信号肽的细胞质定位蛋白,磷酸化修饰主要发生在苏氨酸和丝氨酸位点上,蛋白质的二级结构主要由α-螺旋、无规则卷曲和延伸链组成,STS与查尔酮合成酶(CHS)的蛋白质序列具有高度同源性。4个VdSTS的蛋白质序列在系统进化树上处于3个不同的大分支中,其中,VdSTS5与VdSTS21处在不同的分支中,而VdSTS6与VdSTS20聚在同一分支中,葡萄属不同种之间的STS蛋白质序列相似度高,推测其可能具有相同的功能。启动子顺式作用元件预测发现,4个VdSTS启动子含有多个光响应元件,还含有转录因子识别和作用元件、激素作用元件等。光质显著影响4个VdSTS的表达,在白光、红光、黄光和黑暗处理下,VdSTS的总体表达水平较高,而在绿光和紫光处理下,VdSTS的总体表达水平较低;同时,VdSTS对不同光质的响应不仅存在着基因间的差异,还存在不同培养阶段的差异,VdSTS5在培养后期强烈响应黄光的调控,VdSTS6、VdSTS20在培养中期对黑暗和红光的响应最强,VdSTS21则在培养中期强烈响应红光的调控,这些基因表达水平的变化影响着刺葡萄愈伤组织中白藜芦醇的合成。【结论】4个VdSTS对光质的响应存在差异,可能在响应不同光质调控中对刺葡萄【Objective】The sequence characteristics and expression patterns of stilbene synthases(STS)genes were investigated under different light quality treatments to provide a basis for further analysis on the molecular mechanism of STS-regulated resveratrol synthesis in Vitis davidii(spine grape)callus in response to light signals.【Method】Four STS genes were successfully cloned from V.davidii callus by reverse transcription PCR(RT-PCR).Subsequently,bioinformatic analysis was conducted on these genes and their encoded proteins,and their expression levels were detected in different light qualities and culture stages using quantitative real-time PCR(RT-qPCR).【Result】Four STS genes,VdSTS5,VdSTS6,VdSTS20 and VdSTS21,consisting of 1179 bp complete open reading frame(ORF)and encoding 392 amino acid residues,were successfully cloned from V.davidii callus.These genes were predicted to encode cytoplasm-located hydrophilic proteins and lack signal peptides.Phosphorylation modification mainly occurred at threonine and serine residues,and the secondary structure of proteins were primarily comprised ofα-helices,random coils and extended strands.The protein sequences of STS and chalcone synthase chalcone synthase(CHS)exhibited high homology.The 4 VdSTS proteins were classified into 3 distinct branches in phylogenetic tree,in which VdSTS5 and VdSTS21 were assigned to different branches,while VdSTS6 and VdSTS20 were clustered into a same branch.The STS protein sequences of various Vitis species exhibited high similarity,indicating potential functional conservation.cis-acting element prediction revealed that the promoters of VdSTS contained multiple photoresponsive elements,recognition and binding elements of transcription factor,and hormone regulatory elements.The expression of VdSTS was significantly affected by light quality,with higher overall expression levels under white,red,yellow light and dark treatments,while lower expression under green and purple light treatments.At the same time,the responses of VdSTS to differe

关 键 词：刺葡萄 愈伤组织 STS基因 基因克隆 光质 表达水平 

分 类 号：S663.1[农业科学—果树学] Q786[农业科学—园艺学]