| 三种食源性致病菌多重PCR检测方法的建立及初步应用 |
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| 引用本文： | 沙丹,凌霞,肖勇,吴家林,张敬平. 三种食源性致病菌多重PCR检测方法的建立及初步应用[J]. 上海医学检验杂志, 2009, 0(3) |
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| 作者姓名： | 沙丹 凌霞 肖勇 吴家林 张敬平 |
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| 作者单位： | 无锡市疾病预防控制中心; |
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| 基金项目： | 国家卫生部资助项目(WKJ2006-2-10) |
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| 摘 要： | 目的建立快速检测沙门菌、变形杆菌和金黄色葡萄球菌的多重聚合酶链反应(PCR)方法。方法本研究依据沙门菌侵袭基因正调节蛋白(hilA)基因、变形杆菌溶血素(hpmA)基因和金黄色葡萄球菌特异性pSa-442序列,运用Primer Premier 5.0分别设计3对特异性引物,预计PCR扩增的目的基因片段分别为为580bp、401 bp、256 bp。通过对单管多重PCR扩增的特异性、敏感性分析以及建立L16(43)正交试验对单管多重PCR扩增条件如引物浓度、dNTP浓度和Tm值等的优化,建立了快速同时检测3种食源性致病菌的单管多重PCR方法。结果该方法检测的灵敏度分别为:94.07 pg沙门菌DNA,140.85 ng变形杆菌DNA,1.41 ng金黄色葡萄球菌DNA。模拟检测食品中的混合3种菌,4 h培养后样品的最低检测限度分别为:沙门菌100菌落形成单位(CFU)/mL、变形杆菌101CFU/mL、金黄色葡萄球菌100CFU/mL。结论该方法特异性和灵敏度高,检验周期短,可用于对食品中多种致病菌的快速诊检和监控。
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| 关 键 词： | 多重PCR 沙门菌 变形杆菌 金黄色葡萄球菌 |
Establishment and application of multiplex PCR for diagnosing three food-borne bacterial pathogens |
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| SHA Dan,LING Xia,XIAO Yong,WU Jialin,ZHANG Jingping.. Establishment and application of multiplex PCR for diagnosing three food-borne bacterial pathogens[J]. Shanghai Journal of Medical Laboratory Sciences, 2009, 0(3) |
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| Authors: | SHA Dan LING Xia XIAO Yong WU Jialin ZHANG Jingping. |
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| Affiliation: | Wuxi Center for Disease Prevention and Control;Jiangsu Wuxi 214023;China |
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| Abstract: | Objective To establish a multiplex polymerase chain reaction(PCR) assay for simultaneous detection of Salmonella spp.,Proteus mirabilis and Staphylococcus aureus.Methods Three pairs of primers have been designed according to the hilA of Salmonella spp.,hemolysin(hpmA) of Proteus mirabilis and Staphylococcus aureus clone pSa-442 Sau3AI fragment and the anticipated PCR products were 580bp,401bp and 256bp.A multiplex PCR method has been developed for dignosing Salmonella spp.,Proteus mirabilis and Staphylococc... |
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| Keywords: | Multiplex polymerase chain reaction Salmonella spp. Proteus mirabilis Staphylococcus aureus |
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