首页 > 硕士 > 理学 > 正文

中国香菇自然种质遗传多样性SRAP和IGS2分析

Genetic Diversity in Natural Germplasm of Lentinula Edodes in China by SRAP Marker and IGS2 Analysis

作者: 专业:微生物学 导师:边银丙 年度:2010 学位:硕士  院校: 华中农业大学

Keywords

Lentinula edodes, germplasm, SRAP, IGS2, RFLP, sequence analysis, genetic diversity, variance analysis, agronomic character

        采用SRAP技术对分布在中国9个省份的53个香菇野生菌株进行了遗传多样性分析。选用11对SRAP引物共扩增出491条DNA带,其中95.7%的条带具有多态性,供试菌株在DNA遗传相似性系数上的差异表明中国香菇自然群体遗传多样性丰富,其中横断山脉和云南高原菌株多样性尤其丰富。用平均连锁聚类法构建了样本的遗传相关聚类图,大多数来自同一区域或相邻区域的菌株优先聚成小类,表明菌株的分组与其地理来源明显相关。根据UPGMA聚类结果,以0.77的相似性为切割点,供试菌株可以分为2大类群类群A和类群B,其中A类群还可以分为2个亚类:亚类A1和亚类A2。类群A中,亚类A1主要由9个西北地区和2个东北地区的菌株,以及1个华中地区和1个云南高原的菌株组成;亚类A2主要由西北地区的5个菌株和华中地区的13个菌株组成;类群A还包括2个西北地区的菌株和2个横断山脉的菌株;类群B主要是由分布在云南高原的10个、横断山脉的6个以及台湾的菌株组成。采用IGS2-RFLP技术对53个野生香菇菌株进行了遗传多样性分析,以0.71相似性为切割点,可以将53个菌株分为4个类群,大部分同一地区的菌株聚为一类群。第一类群构成比较复杂,其中包括东北地区的全部菌株,大部分云南地区的菌株,湖北地区的3个菌株,陕西的2个菌株和台湾的菌株;第二个类群由陕、甘、川、云四个地区的菌株组成;第三类群主要由湖北、湖南等地区的菌株组成,还包括个别四川、云南和山西的菌株;第四类群包括湖北和陕西的菌株和1个四川的菌株。对8个菌株的IGS2扩增片段切胶回收克隆并测序,结果表明香菇IGS2序列中的SR2区域由5种亚重复单位组成,亚重复单位又由6种重复元件组成。SR2中亚重复单位的数量不同造成了IGS2长度异质性。对53个野生香菇菌株进行了代料栽培出菇试验,其中14个菌株正常出菇,经方差分析检测农艺性状差异,认为野生菌株之间的平均单袋产量、菌盖直径、菌盖厚度以及平均单菇重的菌株间差异较大。本试验为利用香菇野生资源改良现有栽培菌株提供了重要的试验依据。
    In this study, SRAP markers were utilized to reveal the genetic diversity of 53 strains of Lentinula edodes grown in China, which collected from 9 provinces. A total of 491 DNA bands were detected through 11 primer pairs, of which,95.7% bands were polymorphic. The variations of DNA similarity among the strains reveal that there is great genetic diversity in the natural germplasm of Lentinula edodes in China. According to UPGMA clustering, the 53 strains were classified into 2 groups, and 2 groups also classified into 2 subgroups. Group A1 mainly consisted of strains in the Northwest China and Northeast China; Group A2 mainly consisted of strains in the Northwest China and Central China, while group B comprised the strains of Yunnan Plateau, Hengduanshan mountains and Taiwan. The principal component analysis also classified the strains into 3 groups, support the UPGMA clustering results.Fifty-three wild strains of Lentinula edodes were assessed for DNA polymorphism using IGS2-RFLP technique. According to UPGMA clustering, the 53 strains were classified into 4 groups, Group A1mainly consisted of strains in the Yunnan and Northeast China, group II mainly consisted of strains in the Southwest China, groupⅢcomprised the strains of Central China, while group IV consisted of strains in Hubei and Shanxi.IGS2 fragments of 9 strains were excised from agarose gel, cloned and sequenced. The results displayed that the SR2 was composed of five type of subreapts, and the subreapt was composed of six type of short repeat elements. The heterogeneous lengths of IGS2 are caused mainly by the number of different types of subrepeats within SR2.The fifty-three strains were cultivated with substitute materials on sawdust, and the fruitbodies were successfully produced byl4 strains. The results of variance analysis showed their agronomic characters, expecially for average fruitbody weight, pileus thickness, pileus diameter and average fruitbody weight per bag, are different. This study could be helpful for hybrid between wild and mass production strains.
        

中国香菇自然种质遗传多样性SRAP和IGS2分析

摘要6-7
ABSTRACT7
英文缩略表8-9
1 前言9-18
    1.1 香菇概述9-10
        1.1.1 香菇生活史及其食药用功效9
        1.1.2 香菇遗传育种方法9-10
    1.2 分子标记技术的研究10-14
        1.2.1 RFLP标记11
        1.2.2 RAPD标记11-12
        1.2.3 ISSR标记12
        1.2.4 SSR标记12-13
        1.2.5 AFLP标记13
        1.2.6 SRAP标记13-14
    1.3 DNA序列分析方法——内转录间隔区(ITS)和基因内间隔区(IGS)14-15
    1.4 分子标记在食用菌研究中的应用15-17
        1.4.1 菌株遗传多样性和亲缘关系分析15
        1.4.2 种质资源评价及菌株鉴定15-16
        1.4.3 构建遗传图谱和基因定位16
        1.4.4 核糖体和线粒体遗传16-17
    1.5 本研究的目的和意义17-18
2 材料和方法18-32
    2.1 试验材料18-24
        2.1.1 供试菌株18-19
        2.1.2 供试培养基19-20
        2.1.3 供试试剂20-23
        2.1.4 试验所用仪器设备23-24
    2.2 试验方法24-32
        2.2.1 香菇基因组DNA的提取24
        2.2.2 SRAP反应体系正交设计优化及反应程序24-25
        2.2.3 PAGE电泳及银染操作步骤25-27
        2.2.4 IGS2-PCR、IGS2-RFLP及克隆IGS2片段测序试验方法27-30
        2.2.5 数据分析30-31
        2.2.6 野生香菇菌株出菇试验31-32
3 结果与分析32-53
    3.1 SRAP遗传多样性分析32-40
        3.1.1 香菇基因组DNA的提取32
        3.1.2 SRAP-PCR体系的优化32-33
        3.1.3 反应体系稳定性检测33
        3.1.4 SRAP引物筛选33-34
        3.1.5 DNA扩增产物的多态性34-35
        3.1.6 试验材料的遗传相似性分析35-36
        3.1.7 聚类分析36-40
    3.2 中国香菇野生菌株的IGS2-RFLP分析及IGS2序列分析40-44
        3.2.1 IGS2-PCR体系优化及结果分析40-41
        3.2.2 IGS2-RFLP结果分析41-42
        3.2.3 IGS2测序结果分析42-44
    3.3 野生香菇菌株栽培试验44-53
        3.3.1 栽培袋菌丝生长速度测定44
        3.3.2 转色及现蕾44-45
        3.3.3 农艺性状方差分析45-50
        3.3.4 不出菇菌丝形态及原因50-51
        3.3.5 香菇形态描述与出菇菌株子实体照片51-53
4 讨论53-56
    4.1 中国野生香菇菌株的遗传多样性SRAP分析53-54
    4.2 中国野生香菇菌株IGS2-RFLP遗传多样性及IGS2序列分析54
    4.3 中国野生香菇菌株栽培特性分析54-56
参考文献56-62
致谢62-63
附录 IGS2序列63-71
        下载全文需50


本文地址:

上一篇:恶臭假单胞菌表面展示细菌漆酶及对染料的脱色性能
下一篇:东北天南星凝集素基因的克隆及对蚜虫和褐飞虱的抗性研究

分享到: 分享中国香菇自然种质遗传多样性SRAP和IGS2分析到腾讯微博           收藏
评论排行
公告