应用巢式PCR检测葡萄根癌病菌

摘要

为建立葡萄根癌病菌的分子检测方法,应用细菌核糖体基因间隔片段(16S～23S ribosomal DNA intergenic spacer,ITS)通用扩增引物1405f/456r和葡萄根癌病菌ITS特异引物fI/rI,采用巢式PCR(Nested-PCR)技术对2株葡萄根癌病菌[即葡萄土壤杆菌(Agrobacterium vitis)]和10株非葡萄根癌病菌进行扩增,并且对接种葡萄根癌病菌的匍萄植株进行检测.结果表明,仅有葡萄根癌病菌能扩增出1条长度为607 bp的片段;巢式PCR检测方法可从接种15 d后的葡萄植株中特异性检测出病原菌,而病症的出现需要30～35 d的时间.说明该研究建立的巢式PCR方法可用于快速检测葡萄根癌病菌.%Agrobacterium vitis. which causes crown gall, was detected by nested-PCR using primer pairs 1405f/456r and fl/rl. A band of 607 bp was amplified from two Agrobacterium vitis strains but not from other 10 non-Agrobacterium vitis strains. The pathogenic bacteria could be detected by nested-PCR technique from inoculated individuals at day 15 ,while symptoms could be observed at about day 35. The results of field detection also recommended that nested-PCR technique could be used in quickly earlier detection of crown gall.