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目的建立基于颜色判定的DNA环介导恒温核酸扩增法(loop-mediated isothermal amplification of DNA,LAMP)快速检测金黄色葡萄球菌(SAU)技术。方法利用LAMP法针对SAU特异基因nuc基因设计7套引物,通过对7套引物扩增效率进行比较得到最佳引物组合来快速检测SAU,采用两种结果判断方法在50 min内完成反应。结果 LAMP方法检测核酸的最低浓度为7.16 pg/μl,灵敏度为PCR法的10倍(PCR法为71.6 pg/μl),同时采用22种同源或相似菌种进行特异性检测,结果表明具有良好的特异性。结论 LAMP方法检测过程简单,实验装置简便,反应结果肉眼可视化辨别,灵敏度高,特异性强,特别适合基层防疫与检疫部门对SAU的快速检测。
Objective To establish a rapid detection of Staphylococcus aureus (SAU) by loop-mediated isothermal amplification of DNA (LAMP) based on colorimetric determination. Methods Seven sets of primers were designed according to the nuc gene of SAU specific gene by LAMP method. The optimal primer combinations were obtained by comparing the amplification efficiency of seven sets of primers to detect SAU rapidly. The two methods were used to complete the reaction in 50 min. Results The minimum concentration of nucleic acid detected by LAMP method was 7.16 pg / μl, the sensitivity was 10 times of that of PCR method (71.6 pg / μl by PCR method), and 22 kinds of homology or similar strains were used for specificity test. Specificity. Conclusion The LAMP method has a simple detection process, simple experimental setup, clear visual identification of reaction results, high sensitivity and specificity. It is especially suitable for the rapid detection of SAU by the grass-roots epidemic prevention and quarantine departments.