【作者】 陈庆河； 兰成忠； 陈敦举； 李本金； 翁启勇；

【Author】 CHEN Qing-he LAN Cheng-zhong Chen Dun-ju LI Ben-jin WENG Qi-yong~* (Institute of Plant Protection Fujian Academy of Agricultural Sciences.Fuzhou 350013 China)

【机构】 福建省农业科学院植物保护研究所；

【摘要】 根据辣椒疫霉菌(Phytophthora capsici)和其他疫霉菌及部分卵菌的ras-related protein(Ypt1)基因的序列差异,比较设计了1对特异性PCR引物PCAlF(5‘-GTA TAGCAGAGGTTTAGTGAA-3’)和PCA1R(5‘-ACTGAAGTTCTGCGTGCGTT-3’)。该引物可以从12株来自不同地区的辣椒疫霉菌基因组DNA中特异地扩增出352 bp的片段,而从其他疫霉菌及其他植物病原真菌中均扩增不出条带,表明该引物对辣椒疫霉菌检测具有很好的特异性。进一步优化了检测体系,在25μl的反应体系中,引物PCA1F/PCA1R检测灵敏度为10pg基因组DNA,同时建立了套式PCR检测技术,以PCA1F/PCA1R和PCA2F/PCA2R进行套式PCR扩增,则能够检测到10fg的基因组DNA,检测灵敏度提高了1000倍。更多还原

【Abstract】 A PCR-based method was developed for the identification and detection of Phytophthora capsici in pepper plants.PCR primers PCA1F/PCA1R(PCA1F:5‘-GTATAGCAGAGG TTTAGTGAA-3’, PCA1R:5‘-ACTGAAGTTCTGCGTGCGTT-3’) specific for P.capsici were designed based on the sequence of its ras-related protein(Ypt1)gene.PCA1F/PCA1R amplified a 352 bp product from P.capsici DNA that was not amplified from any other oomycetes or fungi pathogenic.The detection sensitivity with PCA1F/PCA1R was 10pg of genomic DNA in 25μl reaction solution.This sensitivity increased 1000-fold to 10 fg by developing a nested PCR procedure that uses PCA1F/PCA1R as first-round primers combined with PCA2F/PCA2R.更多还原