应用PCR技术快速检测柑桔溃疡病的研究  

Rapid Detection of Citrus Canker Bacteria by Polymerase Chain Reaction

作　　者：唐科志[1] 周常勇[1] 王雪峰[1] 周彦[1] 刘科宏[1] 刘英[1] 

机构地区：[1]中国农业科学院柑桔研究所,重庆400712

出　　处：《中国南方果树》2005年第2期1-4,共4页South China Fruits

基　　金：国家科技攻关"三峡库区特色产业生态经济重建关键技术研究与示范"项目资助

摘　　要：应用PCR检测柑桔溃疡病,所用引物为5'TTGGTG TCGTCGCTTGTAT 3'和5'CACGGGTGCAAAAAATCT3'.从病样的叶片、果皮和枝皮抽提溃疡病菌的PCR扩增,其产物经琼脂粮凝胶电泳,产生大约220bp的特征条带,与纯培养溃疡病菌的PCR产物一致.试验比较两种核酸抽提方法,微量快速抽提法的灵敏度明显高于高盐抽提法.有症状病样品的不同部位均能检测到溃疡病;病株的无症状叶片、果皮、枝皮有部分检测到病菌.病果园健康植株大多数检测不到病菌,少数植株PCR产物电泳务带较弱.取样量以10～20mg为宜.该PCR技术可用于快速检测柑桔溃疡病.A study was conducted for the quick detection of citrus canker by Polymerase Chain Reaction (PCR). The specific primers in DNA amplification are 5' TTG GTG TCG TCG CTT GTA T 3' and 5' CAC GGG TGC AAA AAA TCT 3'. DNA of citrus canker was extracted from infected leaf, fruit peel and stem bark and used as template for the PCR. After PCR and agarose gel electrophoresis, a specific fragment about 220bp has been found in the infected tissue, which is identical with the PCR product from pure-cultured citrus canker bacteria . For DNA extraction, the rapid micro-extraction method is more effective than high salt extraction method. The procedure can be used for the quick detection of citrus canker. The samples taken from the lesion, halo of lesion, tissue nearly the halo and tissue far from lesion in a same leaf, fruit peel or stem bark showed positive result in PCR. A part of the samples taken from leaf, fruit and stem without symptoms in a diseased tree showed positive result. The samples taken from the healty tree in the canker prevailing orchard usually showed negative result.

关 键 词：柑桔溃疡病 溃疡病菌 植株 果皮 病株 叶片 果园 PCR产物 核酸 抽提方法 

分 类 号：S436.66[农业科学—农业昆虫与害虫防治]