西瓜枯萎病菌PCR分子检测方法研究  

Polymerase chain reaction: a molecular diagnostic method for detection of Fusarium oxysporum f.sp.niveum

作　　者：钟鑫 赵静 吴晓蕾[1] 李敬蕊[1] 吕桂云[1] ZHONG Xin;ZHAO Jing;WU Xiaolei;LI Jingrui;Lü Guiyun(College of Horticulture,Agricultural University of Hebei,Baoding 071001,Hebei,China)

机构地区：[1]河北农业大学园艺学院

出　　处：《果树学报》2019年第12期1771-1779,共9页Journal of Fruit Science

基　　金：国家自然科学基金(31872132);河北省自然科学基金(C2016204138)

摘　　要：【目的】西瓜枯萎病是一种严重危害西瓜生产的世界性土传真菌病害,快速及时地检测出西瓜枯萎病菌,对西瓜枯萎病的早期监控预测及后期防治具有重要意义。【方法】利用镰孢属尖孢镰刀菌基因组的数据库,设计筛选西瓜枯萎病菌的特异引物,基于普通PCR技术建立快速简便的分子检测体系,对接种过的西瓜植株和土壤进行检测验证。【结果】该引物可获得556 bp的特异性扩增片段,体系的灵敏度为365 pg·μL^-1。利用PCR检测体系对感病西瓜植株和土壤进行检测,结果显示在抗感品种接种1~5 d后都能检测到西瓜枯萎病菌,而此时植株没有病症,后期检测到的病菌量与其病情指数有一定的相关性;检测接种过的土壤的灵敏度为5×10^2 cfu·g^-1,而在此密度的病菌土壤中,西瓜植株的发病株率较低。【结论】建立了一套基于普通PCR技术分子检测体系,可用于感病植株和土壤的西瓜枯萎病菌快速及时检测,为指导西瓜枯萎病的早期预测以及制定综合防治措施提供重要的参考依据。【Objective】Fusarium wilt of watermelon is a worldwide fungal soil-borne disease caused by Fusarium oxysporum f.sp.niveum(Fon),which is a yield-limiting factor in watermelon production and occurs at all growing stages of watermelon.Like all form special of Fusarium oxysporum,this fungus is a soil inhabitant and extremely difficult to control.Although the use of resistant rootstocks and cultivars provides some degree of protection away from Fusarium wilt,the development of new pathogenic races is a constant problem,and there are currently no commercially acceptable cultivars with adequate resistance to Fon.However,the prevention and control of the disease mainly include crop rotation,grafting and insecticide control,but there are some drawbacks,and the control of watermelon wilt has always been one of the main problems in watermelon production.Therefore,it is very important to establish a simple and rapid early detection method for controlling the disease in watermelon plants and soils.【Methods】Based on the existing genome database of Fusarium oxysporum,the specific primer for qualitative detection of Fon was designed and screened.The PCR detection technology system of Fon was established and optimized.The system was used to detect and verify the inoculated watermelon plants and soils.【Results】The DNA of Fon(race 0,1,2,watermelon),Fusarium oxysporum f.sp.melonis(melon),Fusarium oxysporum f.sp.cucumerinum(cucumber),Fusarium oxysporum f.sp.lycopersici(tomato),Fusarium oxysporum f.sp.conglutinans(cabbage),Fusarium oxysporum f.sp.vesinfectum(cotton)Rhizoctonia Solani(cotton)and Verticillium dahliae(tomato)were used as test materials.Fons(Rcae 0,Rcae 1 and Rcae 2)were all able to amplify the target band with a length of 556 bp using the specific primer FONDX^^-15,while those of other plants were negative.The sensitivity of PCR qualitative detection system was 365 pg·μL^^-1 by using the genomic DNA of Fon as a template.The final reaction system:2×Es TaqMaster Mix 12.5μL,1μL(5μmol·L^^-1)of upstream and

关 键 词：西瓜 枯萎病菌 PCR检测 土壤 

分 类 号：S651[农业科学—果树学]