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由Dickeya spp.引起的香蕉细菌性软腐病是近年来在中国广东发生的一种严重危害香蕉的病害,检测方法的建立和应用是防止病害传播和实时防治的重要手段。依据报道的Dickeyaspp.通用引物,建立了用于香蕉细菌性软腐病发病植株和带菌土壤检测的实时荧光PCR方法。优化后的检测体系对香蕉软腐细菌(XJ8-3-3)靶片段克隆质粒DNA的检测灵敏度可达到2.4×10-5ng·μL-1,对菌悬液的检测灵敏度可达到4.0×102cfu·mL-1,而常规PCR对其检测的灵敏度为2.4×10-3ng·μL-1和4.0×104cfu·mL-1,实时荧光PCR的灵敏度比常规PCR高100倍。利用实时荧光PCR能够快速的检出香蕉细菌性软腐病发病植株和带菌土壤中的病原菌量,对梯度稀释的菌悬液接种的带菌土壤检测结果表明,可检测到病菌DNA最低含量为0.35pg·L-1。该方法适用于对香蕉软腐病菌的检测和监控。
The bacterial soft rot disease of banana caused by Dickeya spp. Is a kind of serious damage to banana in Guangdong in recent years. The establishment and application of detection methods is an important means to prevent disease spread and prevent and cure in real time. Based on the reported Dickeyaspp universal primers, a real-time fluorescence PCR method was established for the detection of banana bacterial soft rot disease and contaminated soil. The optimized detection system could detect 2.4 × 10-5 ng · μL-1 of cloned plasmid DNA of banana soft rot bacterium (XJ8-3-3) target fragment and 4.0 × 102 cfu · mL-1, while the sensitivity of routine PCR was 2.4 × 10-3 ng · μL-1 and 4.0 × 104 cfu · mL-1, respectively. The sensitivity of real-time PCR was 100 times higher than that of conventional PCR. Real-time fluorescence PCR can quickly detect the amount of pathogenic bacteria in banana bacterial soft-rot disease-causing plants and the contaminated soil. The test results on the contaminated soil inoculated with the gradient-diluted bacterial suspension show that the minimum detectable amount of the bacteria DNA is 0.35 pg · L-1. The method is suitable for the detection and monitoring of soft rot of banana.