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首页> 外文期刊>International Journal of Agriculture and Biology >Rapid Identification and Detection of Xanthomonas phaseoli pv. manihotis, Causing Bacterial Blight Disease in Cassava by Real-Time PCR using LNA Probe
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Rapid Identification and Detection of Xanthomonas phaseoli pv. manihotis, Causing Bacterial Blight Disease in Cassava by Real-Time PCR using LNA Probe

机译：快速鉴定和检测黄单胞菌PV。 Manihotis，使用LNA探针通过实时PCR引起木薯中的细菌性枯萎病

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Effective detection and early monitoring of Xanthomonas phaseoli pv. manihotis (Xpm) causing Cassava Bacterial Blight (CBB) disease using rapid DNA-based method is paramount in promoting curative management of the disease. The traditional methods of identification such as pathogen culturing-based upon morphological, biochemical and physiological approaches, traditional PCR involving post-PCR reaction processing gel electrophoresis are more often labour intensive, time-consuming and require exceptional classical taxonomical dexterity. Owing to these limitations of the traditional detection methods, we developed primer and Locked Nucleic Acid probe (LNA probe), and with the aid of Real-time PCR, Xpm was reliably detected from the extracted DNA and directly from the infected cassava plant tissues without DNA extraction within a relatively shorter time (20–25 min). The designed probe has a fluorescent amide (FAM) as reporter dye and carboxytetramethylrodamine (TAMRA) as repressive dye. The selectivity of the developed probe and primer sets were also tested against different Xanthomonas species, and other plant pathogenic bacteria from different genera as well as host cassava genomic DNA. In the end, only Xpm was sensitively and selectively detected by the developed probe, confirming reliable detection and identification of Xpm as the true causal agent of CBB of the infected cassava samples taken from various agro-ecological zones of Ghana. This is the first study, which uses LNA probe, which is mostly used for human pathogens in the identification of cassava plant pathogen.
机译：有效检测和早期监测黄单胞菌PV。使用基于快速DNA的方法导致木薯细菌性疫病（CBB）病的Manihotis（Xpm）在促进该病的治愈管理方面至关重要。传统的鉴定方法（例如基于形态学，生化和生理学方法的病原体培养），涉及PCR后反应处理的凝胶电泳的传统PCR往往是劳动密集型，耗时的，并且需要出色的经典分类灵活性。由于传统检测方法的这些局限性，我们开发了引物和锁定核酸探针（LNA探针），并借助实时PCR技术，从提取的DNA和直接从感染的木薯植物组织中可靠地检测到Xpm，而无需检测在相对较短的时间内（20-25分钟）提取DNA。设计的探针具有荧光酰胺（FAM）作为报告染料和羧四甲基罗丹明（TAMRA）作为抑制染料。还针对不同的黄单胞菌属物种以及来自不同属的其他植物病原细菌以及宿主木薯基因组DNA测试了开发的探针和引物组的选择性。最后，只有Xpm被开发的探针灵敏和选择性地检测到，从而证实了Xpm是从加纳各种农业生态区获取的被感染木薯样品中CBB真正的病因的可靠检测和鉴定。这是第一项使用LNA探针的研究，该探针主要用于鉴定木薯植物病原体的人类病原体。

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