导　　师： 张建农

学科专业： I0202

授予学位： 硕士

作　　者： ;

机构地区： 甘肃农业大学

摘　　要： 甜瓜白粉病是危害我国甜瓜生产的重要病害之一，抗病品种的选育和利用是防治该病害的最有效措施。同时，由于抗病遗传基础的狭窄和生理小种的变异使抗病品种不断丧失抗性，因此，本研究通过甜瓜白粉病病原生理小种鉴定及ssr分子标记寻找抗白粉病紧密连锁标记为出发点，为明确当地白粉病菌的生理小种引进抗源材料，为甜瓜白粉病抗病分子标记辅助育种提高选择效率来开展抗病育种及防治提供理论依据。 本试验对供试甜瓜白粉菌病原贮藏方法、接种方法及生理小种鉴定进行了研究，以抗白粉病品种pmr5和感白粉病品种黄河蜜与其杂交后代f1及f2群体为材料，对亲本、f1和f2单株进行了苗期抗病性接种鉴定，对其进行抗性遗传规律分析，利用f2群体的极抗株和极感株的dna建立抗、感池，用分离群体分组混合分析法（bsa）进行ssr标记分析，寻找与抗病基因连锁的标记，主要实验结果如下： 1．引进国际通用的甜瓜白粉病菌生理小种鉴别寄主与标准鉴定体系，鉴别本试验白粉病菌生理小种，采用不同贮藏方法处理新鲜的白粉病菌，用摩擦接种法和喷雾法将该病原接种于叶片上，观察不同接种方法对白粉病发病的影响，结果显示用阴干低温（4℃）方法贮藏白粉病菌可保持3周左右；摩擦接种法发病早且重；初步确定采自皋兰大棚白粉病菌属单囊壳白粉菌（s．fuliginea）种的生理小种1。 2．构建的prm5×黄河蜜的f2群体性状发生分离，出现抗病和感病植株。田间鉴定123株抗病，57株感病。对f2群体抗感分离数据的x2测验表明，所有f2调查群体抗感分离符合3抗：1感的分离比例（x20．05，1），由此推出pmr5对该病原菌的抗性由1对显性基因控制。 3．ssr稳定性研究中，经过对扩增条件的优化，得出最佳ssr的反应条件为：反应体系（25．0μl）中含有10×taq buffer2．5μl，2．5 mmol/l dntps2．0μl，mgcl2（25 mmol/l）2．0μl，2．5 u/μltaq酶0．4μl，ssr上、下游引物（5pmol/μl）各2．0μl，100 ng/μl模板dna1．0μl。ssr扩增热循环参数为：94℃预变性3 min，然后94℃变性30 sec，50～60℃退火30 sec，72℃延伸1 min，共30个循环，最后72℃延伸5 min。 4．通过分离群体分组分析法（bsa），利用抗病基因池和感病基因池进行抗病基因的ssr标记筛选。经过反复的筛选，引物csat425在亲本和f2代抗感基因池之间扩增出一条大小约为98bp的多态性片段。经f2代群体单株验证后，该多态性条带与pmr5抗白粉病基因连锁，该标记与抗白粉病基因的重组率为13%。定名为csat42598，可以作为甜瓜抗白粉病辅助选择的分子标记。 Powdery mildew of melon which is harmful for melon production is one of the important diseases.It is the most effective measure that the disease-resistant varieties were cultivated and utilized to prevent Powdery Mildew of melon. At the same time,the straitness of genetic basis of disease resistance and variation of physiological races make the disease-resistant varieties continued loss fastness.Therefore, the study is to look for markers that are closely linkage to resistance powdery mildew by SSR molecular markers and Identification of the melon powdery mildew pathogen physiological races which in order to introduce resistance material , to improve selective efficiency using MAS of melon powdery mildew provide theoretical basis for developing breeding and prevetion of disease-resistant varieties. With Parent of resistance PMR5,susceptible resistant Huanghemi and cross-bred F1 and F2 of them as materials,pathogenic Erysiphaceae melon storage methods, vaccination and identification of biological species were studied. Disease-resistant of parents, F1 and F2 plants were inoculated ,identified and analysed genetic regularity in the young plant time. Anti-DNA strain of F2 populations was established to look for linkage marker of disease-resistant gene using Bulked segregant analysis /(BSA/) and SSR markers .The main results showed as follows: 1.International common melon host the standard system of identification of physiological races of powdery mildew,distinguished powdery mildew pathogen physiological races.Fresh powdery mildews were dealed with using different store methods.Using friction and spray inoculation method sprayed the pathogen to leaves,observing disease effect of powdery mildews.The results showed that the dring powdery mildews in low temperature /(4℃/) maintained three weeks; The disease of friction inoculation appeared early and severity powdery mildew collected from greenhouses in Gaolan was initially confirmed S.fuliginea of the Race1. 2.F2 population characters cross bred of PRM5×Huanghemi searated,and resistant and susceptible plants appeared.There were 123 disease-resistant and 57 susceptible by Field identification.χ2 test of resistant separation datas on F2 groups showed that all F2 groups of resistant Separation were in line with 3anti-separation:1 sense of separation /(X20.05, 1/), which concluded that pathogens of PMR5 Resistance was controled by a pair of dominant gene. 3.The best reaction response to SSR are as follows through amplification conditions optimized:The reaction /(25.0μl/) system contained:10×Taq Buffer 2.5μL, 2.5 mmol // L dNTPs 2.0μL,MgCl2 /(25 mmol // L/) 2.0μL,2.5 U //μlTaq enzyme 0.4μL,2.0μL of each primer /(5 pmol //μL/), 100 ng //μL genomic DNA 1.0μL. SSR amplification thermal cycling parameters were : 94℃predegeneration 3 min, then 94℃denaturation 30 sec, 50 ~ 60℃annealing 30 sec, 72℃extension 1 min,there were 30 cycles a total,and at last extended 72℃5 min. 4.Through Bulked segregant analysis /(BSA/),using disease-resistant gene pool and susceptible gene pool filtrated SSR markers of disease resistance gene. After filtrating repeating, primers CSAT425 was amplified approximately 98bp polymorphic fragment between anti-sense gene pool ofparental and that of F2 generation,Polymorphic fragment was linkage to PMR5 powdery mildew resistance gene after F2 plant single was validated ,and the recombination rate of the marker and powdery mildew gene is 13/%.Named CSAT42598 can be used to molecular marker-assisted selection of resistance to powdery mildew of melon.

关 键 词： 甜瓜 抗病品种 抗白粉病基因 病原鉴定 生理小种 标记

分 类 号： [S652]

领　　域： [农业科学] [农业科学]