奶牛乳房炎金黄色葡萄球菌nuc基因LAMP检测方法的建立

Establishment of LAMP Method for Detection of nuc Gene of Staphylococcus aureus from Dairy Cow Mastitis

为了快速鉴定因金黄色葡萄球菌感染引起的奶牛乳房炎,采用环介导等温扩增(LAMP)技术和传统PCR技术,针对金黄色葡萄球菌nuc基因设计特异性引物并进行扩增鉴定,同时对两种鉴定方法的最适反应条件、灵敏度和特异性进行了研究。结果显示,LAMP和传统PCR均能成功扩增金黄色葡萄球菌的nuc基因,但通过优化LAMP反应条件,在62℃时对nuc基因呈现最佳扩增效果,与传统PCR鉴定方法相比,LAMP对金黄色葡萄球菌nuc基因最低检测浓度为50 fg/μL,其检测灵敏度是PCR的10~4倍。说明LAMP能精准鉴定因金黄色葡萄球菌引起的奶牛乳房炎,且针对nuc基因的鉴定操作简单、特异性强、灵敏度高。

In order to quickly identify dairy cow mastitis caused by Staphylococcus aureus infection,loop-mediated isothermal amplification(LAMP)and traditional PCR were used in this study.Specific primers were designed and amplified to identify the nuc gene of Staphylococcus aureus,and studied the optimal reaction conditions,sensitivity and specificity of the two identification methods.The results showed that the nuc gene of Staphylococcus aureus could be successfully amplified by LAMP and PCR.The lowest detection concentration of nuc gene was 50 fg/μL when reaction in LAMP optimal amplification condition(62℃),the detection sensitivity is 104 times than PCR.These results indicated that LAMP was a simple,specific and sensitive method,which could accurately identify dairy mastitis caused by Staphylococcus aureus.