【作者】 赵宏；

【导师】 马峙英；

【作者基本信息】 河北农业大学 ， 作物遗传育种， 2002， 硕士

【摘要】 棉花黄萎病（Verticillium wilt）、枯萎病（Fusarium wilt）是严重危害世界许多国家棉花生产的两大主要病害，也是我国棉花生产上的重要病害。实践证明，利用抗病品种是防治棉花枯、黄萎病的根本措施和有效的途径。 本研究利用AFLP银染技术对30个棉花骨干抗病品种进行了指纹图谱的研究。研究目的包括：第一，建立一套适合棉花指纹图谱分析的AFLP技术体系；第二，获得品种的指纹图谱；第三，结合系谱分析，进行分子水平的遗传多样性研究。主要结果如下： 1．对影响棉花AFLP指纹分析的各个因素进行了试验与摸索。最终建立了一套适合棉花AFLP指纹分析的最适体系。该体系中各优化因素为：①模板DNA浓度为150ng·μL^-1；②酶切体系中，MseI和EcoRI各加入3units，使用NEB buffer #2，反应时间为2h；③连接最适反应时间为10h；④连接产物最适稀释倍数为10倍；⑤预扩增产物最适稀释倍数为10倍。 2．利用AFLP银染技术，采用4碱基切点的MseI和6碱基切点的EcoRI双酶切体系，对30个骨干抗病品种进行AFLP指纹分析。从100个引物组合进行筛选，选出25个引物组合对30个抗病品种进行AFLP分析。25个引物组合共扩出1858条谱带，其中多态性条带数为737条。获得了11个品种的指纹图谱。试验结果证明AFLP银染技术非常适合于棉花遗传多样性及指纹图谱的研究， 3．利用SPSS 10.0软件对AFLPs数据进行聚类分析，得到聚类树状图。多数品种都依据其遗传关系远近进行分类，与系谱资料基本吻合。统计结果结合系谱分析可知，我国现有的陆地棉抗病品种的遗传基础狭窄，遗传变异水平低下，岱字棉系统为主要遗传来源，其次是乌干达棉、斯字棉、金字棉。试验还为两个抗性来源不清的品种进行了分类，表明苏棉4号可能与豫棉一号有相近的亲缘关系；鲁棉12可能与Auburn623RNR有相近的亲缘关系。更多还原

【Abstract】 Fusarium wilt and Verticillium wilt are main diseases which has done harm to the production of cotton in the world, and the same thing also happens in China Practice has proved that it is an effective and important method for preventing Fusarium wilt and Verticillium wilt to grow cultivars with disease resistance.In this paper, AFLP was used to study DNA fingerprinting of 30 main cotton cultivars with resistance to disease. The aim of the study is U)to establish a technical system suitable for fingerprinting study of cotton; (2)to draw DNA fingerprinting of cultivars; (3)to study genetic diversity of cotton combining with pedigree on molecular level. Some results are below:1. At first, some factors affecting establishment of AFLP technological system have been tested and analyzed again and again, and at last a most effective system was established. Some improved factors are below: the concentration of template DNA is 150ng ul; (2)in the restriction system, each of Msel and EcoRI was input 3 units, NEB buffer #2 was used and reaction time is 2 hours; (3)the most available time of ligation reaction is 10 hours;the products of ligation was diluted to 10 times for preamplification;the products of preamplification reaction was diluted to 10 times for selective amplification.2. Using a enzymes combination system including a rare cutter (EcoRl) and a frequent cutter (MseI) to perform AFLP fingerprinting research of 30 cultivars. From 100 primer combines, 25 primer pairs were selected to perform AFLP analysis. 1858 bands were amplified totally and 737 bands were polymorphismic , and 11 cultivars’ DNA fingerprinting were listed. And AFLP technology is very fit for research of genetic diversity in cotton.3. AFLPs were processed by SPSS(Version 10.0). Most cultivars were classified due to their genetic relationship, and results accord to their pedigree. Results also point that two cultivars that missing their genetic origin can be classified: su-mian 4 .cultivars is similar to genetic origin of yu-mian 1; the genetic origin of lu-mian 12 is seem to Auburn623RNR’s.更多还原