| 链球菌、金黄色葡萄球菌、沙门菌和大肠杆菌多重PCR检测方法的建立及应用 |
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| 引用本文： | 许信刚,杨丽花,童德文.链球菌、金黄色葡萄球菌、沙门菌和大肠杆菌多重PCR检测方法的建立及应用[J].中国兽医学报,2012,32(4):534-538,547. |
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| 作者姓名： | 许信刚 杨丽花 童德文 |
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| 作者单位： | 西北农林科技大学动物医学院,陕西杨凌,712100 |
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| 基金项目： | 西北农林科技大学青年学术骨干项目(E111020901);教育部新世纪优秀人才支持计划资助项目(NCET-07-0701) |
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| 摘 要： | 根据NCBI上已收录的链球菌ef-tu基因、金黄色葡萄球菌nuc基因、沙门菌hut基因和大肠杆菌23SrRNA基因的序列,设计并合成4对特异性引物,通过优化多重PCR的反应条件,建立了能够同时检测4种细菌混合感染的多重PCR诊断方法。特异性分析结果表明,应用该方法可以从链球菌、金黄色葡萄球菌、沙门菌和大肠杆菌以及4种细菌的混合物中扩增出4条大小分别为197、278、495和652bp的特异性条带,其他对照组的检测结果均为阴性;敏感性分析表明,该方法对4种病原菌基因组DNA的检出量分别为链球菌25.6pg、金黄色葡萄球菌33.2pg、沙门菌35.7pg、大肠杆菌52.1pg;人工模拟感染样本检测表明,该方法能从混合感染的病料中特异地检测出4种病原菌。本试验建立的多重PCR方法具有特异性强,敏感度高,稳定性好的特点,可以有效的检测链球菌、金黄色葡萄球菌、沙门菌和大肠杆菌的混合感染。
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| 关 键 词： | 链球菌 金黄色葡萄球菌 沙门菌 大肠杆菌 多重PCR |
Establishment and application of multiplex PCR detection method for Staphylococcus aureus,Streptococcus,Salmonella spp and Escherichia coli |
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| XU Xin-gang,YANG Li-hua,TONG De-wen.Establishment and application of multiplex PCR detection method for Staphylococcus aureus,Streptococcus,Salmonella spp and Escherichia coli[J].Chinese Journal of Veterinary Science,2012,32(4):534-538,547. |
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| Authors: | XU Xin-gang YANG Li-hua TONG De-wen |
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| Institution: | (College of Veterinary Medicine,Northwest A&F University,Yangling,Shaanxi 712100,China) |
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| Abstract: | A multiplex PCR assay was developed and evaluated for the clinical detection of Staphylococcus aureus,Streptococcus,Salmonella spp and Escherichia coli.Four pairs of primers were designed and synthesized according to highly conserved regions of the Streptococcus ef-tu gene,Staphylococcus aureus nuc gene,Salmonella spp hut gene and Escherichia coli 23SrRNA gene of NCBI.The multiplex PCR reaction condition was optimized and the multiplex PCR method for detecting the co-infection of Staphylococcus aureus,Streptococcus,Salmonella spp and Escherichia coli was established.The specificity test showed that a fragment of 197,278,495 and 652 bp was amplified from genomic DNA of Streptococcus,Staphylococcus aureus,Salmonella spp and Escherichia coli,respectively.No amplification was achieved from control groups of other bacteria.The sensitivity test showed that the multiplex PCR could detect genome DNA of 25.6 pg for Streptococcus,33.2 pg for Staphylococcus aureus,35.7 pg for Salmonella spp and 52.1pg for Escherichia coli.The artificial simulation test showed that the multiplex PCR could detect four kinds of pathogens from co-infection disease material.The result indicated the multiplex PCR method had advantages of specificity,sensitivity,repetitive,and it could effectively detect the co-infection of Streptococcus,Staphylococcus aureus,Salmonella spp and Escherichia coli in clinical samples. |
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| Keywords: | Streptococcus Staphylococcus aureus Salmonella spp Escherichia coli multiplex PCR |
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