论文部分内容阅读
目的建立快速检测淋球菌porA和16S rRNA基因的环介导等温扩增(Loop-mediated isothermal amplification,LAMP)技术,探讨其用于淋球菌感染早期诊断的可行性。方法利用Primer-ExplorerV3软件分别针对淋球菌porA及16S rRNA基因设计6条引物(2条内引物FIP、BIP,2条外引物F3、B3,2条环引物LF、LB),以淋球菌标准菌株基因组DNA为模板,进行LAMP扩增,同时,以外引物F3、B3为PCR引物,进行PCR扩增,并比较2种扩增方法的灵敏度和特异性。结果LAMP法与PCR法灵敏度相同,可检测到10个拷贝的目的基因;其针对淋球菌porA和16S rRNA基因的引物对脑膜炎奈瑟菌、肺炎链球菌和大肠埃希菌的DNA均不能扩增。结论已成功建立了检测淋球菌porA和16S rRNA基因的LAMP技术,为淋球菌的快速检测提供了新的手段,有望成为淋球菌常规检测的简便方法。
Objective To establish a loop-mediated isothermal amplification (LAMP) technique for rapid detection of porA and 16S rRNA genes in Neisseria gonorrhoeae and to explore its feasibility for early diagnosis of Neisseria gonorrhoeae infection. Methods Primer-ExplorerV3 software was used to design 6 primers (two primers FIP, BIP, two primers F3, B3, two loop primers LF, LB) against gonococcal porA and 16S rRNA genes, Genomic DNA was used as a template for amplification of LAMP. At the same time, primers F3 and B3 were used as PCR primers for PCR amplification. The sensitivity and specificity of the two amplification methods were compared. Results The sensitivity of LAMP method was the same as that of PCR method, and 10 copies of the target gene were detected. The primers of Neisseria gonorrhoeae, Streptococcus pneumoniae and Escherichia coli could not be amplified by primers of porA and 16S rRNA genes increase. Conclusion The LAMP technique for detection of porA and 16S rRNA genes of Neisseria gonorrhoeae has been successfully established, providing a new method for the rapid detection of Neisseria gonorrhoeae. It is expected to be a convenient method for the routine detection of Neisseria gonorrhoeae.