无核白及红地球葡萄VvMADS46基因的克隆及其无核调控功能分析  

Cloning and function analysis of VvMADS46 in Red Globe and Thompson Seedless for seedless regulation

作　　者：胡晓燕 郭春磊 王莉 张松霖 李智[2] 王西平[2] HU Xiaoyan;GUO Chunlei;WANG Li;ZHANG Songlin;LI Zhi;WANG Xiping(College of Horticulture Science and Technology,Hebei Key Laboratory of Horticultural Germplasm Excavation and Innovative Utilization,Hebei Normal University of Science and Technology,Qinhuangdao 066004,Hebei,China;College of Horticulture,State Key Laboratory of Crop Stress Biology in Arid Areas,Northwest A&F University,Yangling 712100,Shaanxi,China)

机构地区：[1]河北科技师范学院园艺科技学院·河北省特色园艺种质挖掘与创新利用重点实验室,河北秦皇岛066004 [2]西北农林科技大学园艺学院·旱区作物逆境生物学国家重点实验室,陕西杨凌712100

出　　处：《果树学报》2021年第8期1231-1239,共9页Journal of Fruit Science

基　　金：国家自然科学基金-联合基金重点项目(U1603234)。

摘　　要：【目的】探究VvMADS46与葡萄无核性状产生的关系。【方法】采用实时荧光定量PCR分析VvMADS46在葡萄不同组织器官的表达,分析克隆VvMADS46基因序列,构建系统发育树,进行VvMADS46基因的亚细胞定位分析及过量表达番茄性状观察,研究其功能。【结果】VvMADS46在无核白和红地球的根、茎、叶、花序、花、卷须、果实等不同组织中均有表达,在花中的表达量较高,在果实中的表达量次之,而在茎、叶和卷须中的表达量较低;花后33、36、39、42 d 4个时期无核葡萄无核白、火焰无核VvMADS46的表达量都显著高于有核葡萄红地球、巨峰;构建了VvMADS46基因的亚细胞定位载体,通过农杆菌侵染洋葱,发现了VvMADS46定位在细胞核上;克隆了VvMADS46基因,在NCBI数据库进行序列比对,得到其同源基因为AP3基因,筛选得到AtAP3、AeAP3、CmAP3、LjAP3、LrAP3、PhAP3、AcAP3和PtAP3,并构建了系统发育树;构建了VvMADS46基因的植物过量表达载体,采用农杆菌介导法转化模式植物番茄,发现转基因番茄植株明显矮小且种子显著变小。【结论】VvMADS46与葡萄胚珠败育、无核性状发生存在密切联系。【Objective】Grapevine(Vitis vinifera L.)as one of the most economically important fruit crop is widely cultivated all over the world.Seedlessness of the fruit is one of the most desired traits for table grape consumers.The MADS-box genes,which generally comprise large families,encoded transcription factors(TFs)with a highly conserved MADS domain,and played diverse roles in plant embryo development,flowering time,floral meristem,and fruit ripening regulation.In this study,VvMADS46 was expressed in the different grape cultivated species tissues and ovule developmental stages.Gene cloning,sequence alignment and overexpression in the Micro-Tom tomato were analyzed for VvMADS46 gene related grape seedless function identification,in order to provide foundation for further research of VvMADS46 participating in regulation of seedless grapes.【Methods】Grape tissues and organs,including roots,stems,leaves,tendrils,inflorescences,flowers in the fully opening stage and fruits on 33 days after flowering(DAF)were harvested from the seedless grape Thompson Seedless and the seeded grape Red Globe,respectively.Ovules were collected from the two seeded grape cultivars(Kyoho and Red Globe)and the two seedless grape cultivars(Thompson Seedless and Flame Seedless)on 33,36,39 and 42 DAF.Total RNA extraction and first strand cDNA synthesis were accomplished using EZNA Plant RNA Kit(R6827-01,Omega Bio-tek,USA)and PrimeScript 1st Strand cDNA Synthesis Kit(TaKaRa Biotechnology,Dalian,China),respectively.VvMADS46 expression in the different grape tissues and organs was made by quantitative real-time PCR.VvMADS46 cDNA sequences,alignment and phylogenetic tree were analyzed by homologous clone.The sub-cellular localization and the overexpression of VvMADS46 were analyzed in the onion and the model plant Micro-Tom tomato.【Results】VvMADS46 was expressed in all the different tissues and organs of the Thompson Seedless and the Red Globe.Obviously,there was no significant difference in the roots,stems,leaves and tendrils between the see

关 键 词：葡萄 VvMADS46 基因克隆 无核调控 

分 类 号：S663.1[农业科学—果树学]