大麦黄矮病毒运动蛋白与AtVOZ1和AtVOZ2互作并延迟拟南芥开花

发布时间：2018-01-10 09:33

本文关键词：大麦黄矮病毒运动蛋白与AtVOZ1和AtVOZ2互作并延迟拟南芥开花　出处：《西北农林科技大学》2017年硕士论文　论文类型：学位论文

【摘要】：大麦黄矮病毒(Barely yellow dwarf viruses,BYDVs)侵染多种禾本科作物,呈现多种发病症状同时也给世界粮食生产造成严重的损失。其在小麦上的症状主要有植物矮化、叶片黄化,除此之外在燕麦上还能造成分蘖和抽穗的减少。而目前对于这种症状的产生原因研究的还不是很清楚。为了更好地控制小麦病毒病发生,研究该病症的发病机制势在必行。大麦黄矮病毒的运动蛋白(Movement Protein,MP)的主要功能是结合病毒RNA,增大胞间连丝孔径并促使其在植物细胞间运输。但是对于MP对植物生理生化方面的影响研究的还是较少。因此,本研究针对BYDV-GAV MP对模式植物拟南芥生理生化方面的影响进行了研究,主要结果如下:1.通过酵母双杂交实验,发现在拟南芥中能够与MP蛋白互作的是两个转录因子,维管植物锌指蛋白AtVOZ1和AtVOZ2。利用双分子荧光互补实验(BiFC),进一步验证MP与AtVOZ1和AtVOZ2是能够相互作用的,且互作的位置是细胞质。根据MP蛋白的关键结构域,对MP片段进行缺失突变,构建了MP(DH1)、MP(DH2)、MP(DH3)三个突变体。通过双分子荧光互补实验,结果表明只有MP(DH2)不能与AtVOZ1和AtVOZ2互作,而MP(DH2)是缺失了MP的N端1-81aa,即MP的核定位信号区域,说明MP的N端核定位信号在互作中是必须的。2.由于AtVOZ1和AtVOZ2是控制拟南芥开花通路上的关键基因。我们通过农杆菌浸花转序的方法构建MP转基因拟南芥。探究转基因植株是否产生开花方面的表型差异。结果显示,同时播种野生型和转基因拟南芥,开花时,转基因拟南芥莲座叶数目比野生型显著增多,同时转基因拟南芥经过更长的时间才开始开花。MP转基因拟南芥表现晚花表型,也就是MP影响了拟南芥的开花表型。3.AtVOZ1和AtVOZ2是光周期通路phyB途径上的关键基因。为了探究MP转基因植株出现的晚花表型是否与phyB途径有关,通过实时荧光定量PCR试验,测定转基因拟南芥及野生型拟南芥phyB通路上的关键基因AtVOZ1、AtVOZ2、CO、FLC、FT的表达量。结果表明MP转基因植株的可能通过调节FT基因的表达量来影响拟南芥开花。综上所述,本实验揭示了BYDV-GAV MP通过与AtVOZ1、AtVOZ2的互作,可能通过影响FT基因的表达,介导晚花表型。为BYDV-GAV对症状的产生机理研究奠定基础。
[Abstract]:Barley yellow dwarf virus (Barely yellow dwarf viruses, BYDVs) infection of a variety of gramineous crops, showing a variety of symptoms but also to the world food production caused serious losses. The main symptoms in wheat plant dwarfing, Leaf Chlorosis, in addition to the decrease in oat also can cause tillering and heading and the current. For this reason of symptoms is not very clear. In order to control wheat virus disease, it is imperative to study the pathogenesis of disease. The movement protein of barley yellow dwarf virus (Movement Protein MP) is the main function of virus RNA, enlarged plasmodesmata aperture and the transport in plant cells. But for MP on plant physiological and biochemical aspects of the impact of research is still less. Therefore, this study was about the influence of BYDV-GAV MP on Arabidopsis physiological and biochemical aspects. The main research results are as follows: 1. by the yeast two hybrid experiment, found to interact with MP protein in Arabidopsis is two transcription factor, vascular plant zinc finger protein AtVOZ1 and AtVOZ2. using bimolecular fluorescence complementation assay (BiFC), MP and AtVOZ1 and further validation of AtVOZ2 is able to interact with each other. The position is the key. According to the cytoplasmic domain of MP protein, the MP deletion mutation, constructed MP (DH1), MP (DH2), MP (DH3) three mutants by bimolecular fluorescence complementation experiments, the results show that only MP (DH2) with AtVOZ1 and AtVOZ2 interaction, and MP (DH2) is the lack of N end 1-81aa MP, namely the nuclear localization signal region of MP, N MP in the end of the nuclear localization signal interaction is necessary because the AtVOZ1 and.2. AtVOZ2 is the key gene control flowering pathway. By Agroinoculation method to order the bacteria leaching flower building MP transfer matrix Because of the Arabidopsis transgenic plants. To explore whether phenotypic differences in flowering. The results showed that the sowing and wild type and transgenic Arabidopsis, transgenic Arabidopsis rosette leaves of flowering, the number increased significantly than that of the wild type, the transgenic Arabidopsis plants after a longer period of time to flowering.MP transgenic Arabidopsis showed late flowering phenotype, which is the MP effect in Arabidopsis thaliana the flowering phenotype of.3.AtVOZ1 and AtVOZ2 are the key genes in the photoperiod pathway phyB pathway. In order to explore the MP transgenic plants appeared late flowering phenotype is associated with phyB pathway by real-time fluorescence quantitative PCR test, determination of key gene AtVOZ1 in transgenic Arabidopsis and wild type Arabidopsis phyB pathway on AtVOZ2, CO, FLC, FT expression MP. The results showed that transgenic plants may be through the regulation of FT gene expression to affect the flowering of Arabidopsis. In summary, this study reveals The interaction between BYDV-GAV MP and AtVOZ1 and AtVOZ2 may mediate the late flowering phenotype by influencing the expression of FT gene, and lay the foundation for the study of the mechanism of symptom generation by BYDV-GAV.

【学位授予单位】：西北农林科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S432.41

【相似文献】