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应用复合PCR方法检测沙门菌的研究

吴华， 高宏伟， 朱来华， 徐彪， 王树峰， 梁成珠， 毕建秀

（青岛出入境检验检疫局， 山东 青岛266002）

摘要： 参照文献报道的沙门菌Repeat se和hisJ基因片段的引物序列，设计并合成了2对引物，对沙门菌属和非沙门菌属的标准菌株及分离菌株进行了复合PCR扩增反应。结果，沙门菌PCR产物均出现199bp与495bp的特异性DNA扩增条带，而非沙门菌均未出现扩增条带，证明这2对引物具有沙门菌属特异性。将提取的沙门菌DNA做梯度稀释，测定该复合PCR体系的敏感度。结果表明，此体系可检出103CFU/mL菌液提取的DNA模板。应用上述方法，对进境鱼粉阳性样品进行了检测，均出现阳性结果。本研究表明，复合PCR是一种特异、敏感、快速的沙门菌检测方法，可应用于口岸系统鱼粉、肉骨粉的日常检测。

关键词： 复合PCR； 沙门菌； 检测

中图分类号：S 852.612 文献标识码： A  文章编号：1000-6419(2003)12-0015-05

Study on detection of Salmonella by multiplex PCR

WU Hua，GAO Hongwei，ZHU Laihua， XU Biao， WANG Shufeng， LIANG Chengzhu， BI Jianxiu

(Qingdao EntryExit Inspection and Quarantine Bureau of the People’s Republic of China, Qingdao 266002, China)

Abstract: A multiplex PCR method was established for detection of Salmonella by using two pairs of primers unique to Repeat sequence and hisJ gene segments of Salmonella species. Two specific DNA amplicons were present in the standard strains and laboratory isolates of Salmonella, but absent in the nonSalmonella samples, which showed that two pairs of primers are Salmonellaspecific. The sensitivity of this method was confirmed by serial dilutions of extracted SalmonellaDNAs while approximately 103 colony forming units per 1mL (CFU/mL) were detected. Moreover, positive samples, indicated by conventionalmicrobiological assays followed by confirmative serological tests in about seven days, were also demonstrated by the multiplex PCR within hours. It is revealed in this study that the multiplex PCR, specific, sensitive, rapid, can be applied to routine test of imported fish meal, meat and bone meal.

Key words: multiplex PCR; Salmonella; detection

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