摘要
根据梨火疫病菌16S^23S间的ITS保守序列,设计并合成了一对特异性引物REA/FEA,应用荧光染料SYBRG reen I,对10个梨火疫的菌株和其它相关参试菌株进行了检测。结果表明,10个梨火疫菌株都产生荧光信号而其它参试菌株都不产生荧光信号,成功建立了梨火疫病菌的实时荧光PCR检测方法。整个检测过程只需3 h,完全闭管,降低了污染的机会,无需PCR后处理。检测的灵敏度是4个菌体细胞,比常规PCR电泳检测提高了10倍。用该特异性引物对梨枝条浸泡液进行实时荧光PCR检测,结果可特异性检测到目标菌的存在,并且检测的灵敏度是24个菌体细胞,比常规PCR电泳检测提高10倍。
One pair of specific primes （REA/FEA） is designed based on the conserved sequence of internal transcribe spacer （ITS）between 16S rDNA and 23S rDNA of E. amylovora,SYBR Green I real-time fluorescent PCR method, which has no post-PCR manipulations and reduces false positive result and contamination risks, is established for detection of this bacteritum. In all tested bacterial strains, only E. amylovora can be detected. Detection limitation of pure culture and artificially inoculated samples were 4 and 24 bacterial cells, sensitivity was 10 times higher than PCR gel electrophoresis detection.
出处
《植物病理学报》
CAS
CSCD
北大核心
2006年第2期123-128,共6页
Acta Phytopathologica Sinica
基金
国家重点基础研究发展规划项目(2002CB111400)
国家高技术研究发展计划项目(2003AA249020)
