黑龙江省稻瘟病菌致病性与无毒基因检测及遗传多样性分析

发布时间：2018-05-06 11:34

本文选题：稻瘟病菌 + 致病性　；参考：《黑龙江八一农垦大学》2017年硕士论文

【摘要】：本研究利用2014和2015年在黑龙江省30个市(县)采集的稻瘟病菌穗颈瘟标样,获得稻瘟病菌的单孢菌株,对其进行对主栽品种的致病能力、无毒基因在各地区的分布情况和群体结构的研究,明确了黑龙江省稻瘟病菌群体的多样性和变异的复杂性,为黑龙江省水稻种植的合理布局、抗病种质资源的筛选和培育提供理论依据。(1)以苗期接种病原菌的方法,以致病率、毒力频率和联合致病系数为指标分析171个稻瘟病菌对18个水稻主栽品种的致病性。结果表明:不同的稻瘟病菌对供试主栽品种的致病能力有明显差异,致病率范围从0~78%;供试菌株对各供试品种的毒力率明显不同,龙粳31的毒力频率最低,为1.17%;以联合致病性为指标探究不同品种组合与稻瘟病菌间的互作,龙洋16+龙粳31、龙粳31+龙粳43、龙粳31+松粳20为高抗的搭配组合,其RAC大于0.8,VAC为0,可以在生产推广中搭配使用。(2)采用设计特异性引物的方法对288个稻瘟病菌菌株进行PCR扩增,检测稻瘟病菌无毒基因AVRPiz-t和AVR-Pita的存在情况并对部分菌株进行测序分析。结果表明:无毒基因AVRPiz-t和AVR-Pita在黑龙江省的出现频率分别为56.6%和22.9%。2014年和2015年无毒基因AVRPiz-t的出现频率分别为37.0%和95.8%,AVR-Pita的基因频率分别为23.0%和22.9%。供试菌株的无毒基因在年份、地区、基因之间都存在明显差异。对部分菌株的测序分析发现两个无毒基因在黑龙江省稻瘟病菌群体中都存在变异的情况,特别是AVR-Pita存在多个变异位点和类型,这些改变可能会导致基因功能缺失,失去无毒性。表明黑龙江省稻瘟病菌株变异的易变性和遗传多样性。(3)采用Pot2-rep-PCR指纹识别方法对227个菌株进行扩增,研究黑龙江省菌株的遗传多样性。结果表明:供试菌株分别扩增到2~13条DNA条带,大小在500~6000 bp之间。经UPGMA聚类分析,2014年的162个菌株在0.73的遗传距离下被划分为10个遗传谱系。2015年65个菌株在0.77的遗传距离下被划分为8个遗传谱系。2014和2015年共227个稻瘟病菌菌株在0.75的相似水平下被划分为12个遗传宗谱。黑龙江省稻瘟病菌在地区和年份之间遗传谱系都有很大的变化,说明了黑龙江省稻瘟病菌的高度变异和丰富的遗传多样性。
[Abstract]:In this study, Monosporium strains were collected from 30 cities (counties) of Heilongjiang Province in 2014 and 2015, and their pathogenicity to the main cultivars was studied. The study on the distribution and population structure of nontoxic genes in different regions made clear the diversity and variation complexity of rice blast fungus in Heilongjiang Province, which was the rational distribution of rice cultivation in Heilongjiang Province. The selection and cultivation of resistant germplasm resources provided theoretical basis for the selection and cultivation of disease-resistant germplasm resources. The pathogenicity of 171 rice blast germplasms to 18 main rice cultivars was analyzed by inoculating pathogenic bacteria at seedling stage with the index of pathogenicity, virulence frequency and combined pathogenicity coefficient. The results showed that the pathogenicity of different strains of rice blast was significantly different from that of 0.788.The virulence rate of the tested strains was obviously different, and the frequency of virulence of Longjing 31 was the lowest. To explore the interaction between different combinations and rice blast fungus, Longyang 16 longjing 31 longjing 43 long japonica 31 Songjing 20 was used as high resistance combinations. Its RAC is 0, which can be used in production and popularization. The designed specific primers were used to amplify 288 strains of Magnaporthe grisea by PCR. The presence of non-toxic genes AVRPiz-t and AVR-Pita of Magnaporthe oryzae was detected and some strains were sequenced. The results showed that the frequencies of nontoxic gene AVRPiz-t and AVR-Pita in Heilongjiang Province were 56.6% and 22.9% in 2014 and 2015, respectively, which were 37.0% and 95.8%, respectively. The frequencies of AVR-Pita were 23.0% and 22.9%, respectively. The nontoxic genes of the tested strains were significantly different in year, region and gene. Sequencing analysis of some strains showed that the two nontoxic genes were mutated in the population of blast fungus in Heilongjiang Province, especially there were many mutation sites and types in AVR-Pita. These changes may lead to the deletion of gene function and the loss of nontoxicity. The results showed that the variability and genetic diversity of blast strains in Heilongjiang Province were analyzed by Pot2-rep-PCR fingerprinting method to study the genetic diversity of Heilongjiang Province strains. The results showed that 213 DNA bands were amplified from the tested strain, and the size was between 500,000bp and 6000bp. Based on UPGMA cluster analysis, 162 strains in 2014 were divided into 10 lineages at 0.73 genetic distance. 65 strains were divided into 8 lineages at 0.77 genetic distance in 2015. A total of 227 strains of Magnaporthe grisea were divided into 0.75 in 2014 and 2015. The genetic genealogy was divided into 12 genetic genealogies at similar levels. The genetic pedigree of rice blast fungus in Heilongjiang Province varied greatly between regions and years, which indicated the high variation and rich genetic diversity of rice blast fungus in Heilongjiang Province.
【学位授予单位】：黑龙江八一农垦大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S435.111.41

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