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目的建立聚合酶链反应(PCR)方法快速检测牛奶、冰淇淋及肉中的金黄色葡萄球菌(金葡菌)。方法针对金葡菌耐热核酸酶基因(nuc)、纤维蛋白原结合蛋白基因(ClfA)设计并合成2对引物进行PCR扩增,特异的检测金葡菌,并对52株金葡菌和31株非金葡菌进行扩增,验证方法的特异性。在牛奶、冰淇淋及肉中人工污染不同菌量,增菌培养,在不同时间段提取DNA进行PCR扩增,确定该方法在食品中的检出限。结果nuc、ClfA两对引物对金葡菌的检出均有很好的特异性,在3种食品中的检出限均为10cfu/g(ml),全部检测可在24h完成。结论建立了适用于牛奶、冰淇淋及肉中的金葡菌检测的快速、灵敏、特异的PCR方法。
Abstract：Objective To establish a rapid polymerase chain reaction (PCR)method for detection of staphylococcus aureus in milk, ice cream and meat. Methods Two pairs of oligonucleotide primers were designed with thermo nuclease gene nuc and surfaced-associated fibrinogen-binding protein gene ClfA to detect staphylococcus aureus. Fifty-two staphylococcus aureus and thirty-one non staphylococcus aureus were amplified by PCR to verify the specificity. Various numbers of bacteria were added into milk, ice cream and meat. After enrichment, DNA extracted in different time was amplified by PCR to verify detection limit. Results Each primer pair allows specific detection. The limit of detection was 10cfu/g(ml) in three kinds of food. Whole procedure of detection could be finished in 24 hours. Conclusion A rapid, sensitive and specific PCR method can be applied detecting staphylococcus aureus in milk, ice cream and meat.
1ATANASSOVA V,MEINDL A,RING.Prevalence ofStaphylococcus aureusand staphylococcal enterotoxins in rawpork and uncooked smoked ham-a comparison of classical culturing detection and RFLP-PCR[J].Int J Food Microbiol,2001,68:105-113.
2陈靖,陈叶.用酶联免疫吸附法(ELISA)检测金黄色葡萄球菌肠毒素[J].中国食用菌,1999,18(5):39-40.
3OMOE K,ISHIKAWA M,SHIMODA Y.Detection of seg,seh and sei genes instaphylococcus aureusisolates and determination of the enterotoxin productiveities of S.aureus isolates harboring seg,seh or sei genes[J].J Clin Microbiol,2002,40(3):857-862.
4YAZDANKHAHSP,SOLVEROD L,SIMONSEN S.Development and evaluation of an immunomagnetic separation-ELISAfor the detection of staphylococcus aureusthermostable nuclease in composite milk[J].Vet Microbiol,1999,67(2):113-125.
5WILSONL G,COOPERJ E,GILMOUR A.Detection of enterotoxigenic staphylococcus aureusin driedskimmed milk:use of the polymerase chain reaction for amplification and detection of staphylococcal enterotoxin genes entB and entC1andthe thermonuclearse gene nuc[J].Appl and Environ Microbiol,1991,57(6):1793-1798.
6HOOKEYJ V,RICHARDSONJ F,COOKSONB D.Moleculartypingof staphylococcus aureusbased on PCR restriction fragment length polymorphismand DNAsequence analysis of the coagulase gene[J].J Clin Microbiol,1998,36:1083-1089.
7MCDEVITT D,NANAVATY T,HOUSE-POMPEO K,et al.Charact-erization of the interaction between theStaphylococcus aureusclumping factor(ClfA)and fibrinogen[J].Eur J Biochem,1997,247(1):416-424.
8SMELTZER MS,GILLASPY AF,PRATTF L,et al.Prevalence and chromosomal maplocation of staphylococcus aureus adh-esion genes[J].Gene,1997,196:249-259.
基本信息:
中图分类号:R155.5
引用信息:
[1]田静,计融,杨军,等.PCR方法快速检测食品中的金黄色葡萄球菌[J].卫生研究,2007(02):183-186.
基金信息:
国家科技部十五攻关重大项目(No2001BA804A03)
2007-03-30
2007-03-30