山东和安徽地区三种蔬菜病毒检测与鉴定

发布时间：2018-04-16 04:30

本文选题：番茄曲叶病毒 + 番茄褪绿病毒　；参考：《山东农业大学》2016年硕士论文

【摘要】：番茄及辣椒都是重要的经济作物,近年来病毒病的爆发导致了番茄和辣椒产量减少和品质的降低,造成了严重的经济损失。随着各种检测手段的发展,我们可以更迅速、更便捷的鉴定病毒病的种类,并采取相关措施挽回经济损失。本研究以番茄黄花曲叶病毒(Tomato yellow leaf curl virus, TYLCV)、番茄褪绿病毒(Tomato chlorosis virus, ToCV)及辣椒轻斑驳病毒(Pepper mild mottle virus, PMMoV)为对象,用分子检测方法完成山东、安徽两省病原的鉴定。我们选取山东、安徽两个省7个地区,针对3种病毒病进行田间调查,采集疑似样品扩增序列,对其进行同源性比对和遗传进化树等分析,进一步了解山东、安徽两省蔬菜产区3种病毒病的发生情况。主要研究结果如下：1、通过对山东省、安徽省TYLCV发生情况的调查及检测可知,目前TYLCV在山东省大部及安徽省局部已经发生扩散。构建的TYLCV系统进化树可以分为IL、IR、 Mid、Gez4组,其中本研究的4个分离物和中国所有的分离物一起聚类在IL组。对感病TYLCV样品进行卫星DNA的检测,没有获得明显目的条带。2、通过RT-PCR检测及序列分析,首次确定ToCV在安徽地区发生,且统计发现两省六个地区ToCV、TYLCV存在单独侵染及复合侵染。分别基于ToCV CP和ToCV HSP70构建系统进化树,结果发现ToCV分离物都可分为两个大组,其中本研究所得到的分离物均与中国其他地区的分离物聚类到Ⅰ组。通过计算分离物ToCV的CP与HSP70两个基因的同义突变(dS)和非同义突变(dN)的比率(dN/dS),结果显示,CP与HSP70基因均处于负向(或净化)选择的压力之下且所承受的压力不同。3、通过对山东济宁辣椒的田间调查发现,PMMoV在辣椒叶片上呈现轻微的褪绿或者斑驳症状。对山东济宁辣椒病株进行RT-PCR扩增,经鉴定,确定其病原为PMMoV,表明感病的济宁辣椒样品确系被PMMoV感染。并与日本分离物(AB062054)、广西分离物(KP877405)2个序列同源性最高均达到99.2%。基于CP构建的系统进化树发现济宁分离物与中国其他分离物聚类到Ⅰ组,与中国北京分离物(PMMoV-CN)最为相近。
[Abstract]:Tomato and capsicum are important cash crops. In recent years, the outbreak of virus disease has led to the decrease of tomato and pepper yield and quality, resulting in serious economic losses.With the development of various detection methods, we can identify the virus disease more quickly and conveniently, and take relevant measures to recover the economic loss.In this study, Tomato yellow yellow leaf curl virus, TYLCV, Tomato chlorosis virusand Pepper mild mottle virus. were used to identify the pathogens in Shandong and Anhui provinces by molecular detection.We selected 7 regions of Shandong and Anhui provinces to carry out field investigation on three kinds of virus diseases, collected suspected sample amplification sequences, analyzed their homology comparison and genetic evolution tree, and further understood Shandong Province.Occurrence of three kinds of virus diseases in vegetable production areas of two provinces in Anhui Province.The main results are as follows: 1. By investigating the occurrence of TYLCV in Shandong Province and Anhui Province, we can see that TYLCV has spread in most of Shandong Province and Anhui Province at present.The TYLCV phylogenetic tree can be divided into 4 groups, I. e., ILR, Midge, and Gez. The four isolates in this study are clustered together with all the Chinese isolates in the IL group.The detection of susceptible TYLCV samples by satellite DNA showed that there was no obvious target band. By RT-PCR detection and sequence analysis, it was determined for the first time that ToCV occurred in Anhui area, and it was found that there was single infection and compound infection in six regions of two provinces.Phylogenetic tree was constructed based on ToCV CP and ToCV HSP70. The results showed that the ToCV isolates could be divided into two groups.By calculating the ratio of CP and HSP70 gene synonymous mutation (DS) and non-synonymous mutagenesis (DN) of isolate ToCV, the results show that both CP and HSP70 genes are under negative (or purifying) selective pressure and the pressure is different.A field investigation of Jining pepper in Shandong Province showed that PMMoV showed slight chlorotic or mottled symptoms on pepper leaves.The pathogen of Jining pepper was confirmed as PMMoV by RT-PCR amplification, which indicated that the susceptible pepper sample was infected by PMMoV.The highest homology was found between the two isolates, AB062054 and KP877405, respectively, and the highest homology was 99.2% with the Japanese isolate AB062054 and the Guangxi isolate KP877405.The phylogenetic tree constructed based on CP showed that Jining isolates were clustered to group I with other Chinese isolates, which was most similar to PMMoV-CNS isolated from Beijing, China.
【学位授予单位】：山东农业大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：S436.3

【相似文献】