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为了提高口岸和基层实验室检疫和监测玉米细菌性枯萎病菌的准确性和工作效率,利用环介导恒温扩增技术(LAMP),根据内切葡聚糖酶(EGase)基因前导序列,设计2个内引物和2个外引物,对玉米细菌性枯萎病菌进行快速检测。结果表明,使用玉米细菌性枯萎病菌的近缘种或引致相似症状的病原菌菊欧文氏菌玉米致病变种Erwinia chrysanthemi pv.zeae、玉米内州萎蔫病菌Clavibacter michiganensis subsp.nebraskensis、燕麦假单胞菌Pseudomonas avenae、杓兰欧文氏菌Erwinia cypripedii检测其特异性,仅玉米细菌性枯萎病菌有扩增。LAMP检测灵敏度达到2 pg DNA,为普通PCR的100倍;与其它检测方法相比,LAMP方法检测时间短,效率高,不仅降低了设备投入,易于操作,而且具有较高的灵敏度和特异性,适合玉米细菌性枯萎病菌的现场检疫和大规模检测。
In order to improve the accuracy and efficiency of quarantine and monitoring corn bacterial wilt pathogens in port and grass-roots laboratories, using ring-mediated isothermal amplification (LAMP), according to the leader sequence of endoglucanase (EGase), design 2 One internal primer and two external primers for rapid detection of bacterial wilt pathogen of corn. The results showed that Erwinia chrysanthemi pv. Zeae, Clavibacter michiganensis subsp. Nebraskensis, Pseudomonas sp. Strains, which were caused by relatives of pathogenic bacteria of pathogenic bacteria Fusarium oxysporum, avenae and Erwinia cypripedii, respectively. Only the bacterial wilt pathogen of corn was amplified. LAMP detection sensitivity of 2 pg DNA, 100 times for the normal PCR; Compared with other detection methods, LAMP detection time is short, high efficiency, not only reduces the equipment investment, easy to operate, but also has high sensitivity and specificity, Suitable for on-site quarantine and large-scale testing of corn bacterial wilt pathogens.