摘要
目的应用环介导等温扩增(loop-mediated isothermal amplification,LAMP)技术建立一种快速敏感的脑膜炎奈瑟菌属检测方法。方法对脑膜炎奈瑟菌种属基因(ctr A)序列的6个区域设计4条LAMP引物(2条内引物、2条外引物),同时设计2条环引物,并对反应条件和反应体系进行优化。分别验证该方法的特异性及敏感性,并与PCR检测方法进行比较。结果适宜反应条件所设计引物对脑膜炎奈瑟菌采用LAMP扩增技术,其特异性及敏感性均高于PCR检测。结论本实验建立的LAMP方法能够快速、灵敏、特异地检测脑膜炎奈瑟菌。相比普通PCR更快速,在60 min内即可完成扩增反应,扩增效率高,反应不需要精密控温设备和高级复杂的分析仪器,对操作人员的熟练度和专业水平要求不高,适合基层检验部门及小型实验室与现场监测等使用。
Objective To establish a rapid sensitive detection approach for Neisseria meningitids（ Nm） based on the loop-mediated isothermal amplification（ LAMP） technique. Methods Four LAMP primers（ two inner and two outer） were designed according to the six zones of the ctr A gene of Nm,and two ring primers were designed and the reaction conditions and system were optimized. The sensitivity and specificity of the detection approach were evaluated and compared to conventional PCR methods. Results The primers designed for suitable reaction conditions for Neisseria meningitids were amplified by LAMP,and their specificity and sensitivity were higher than those detected by PCR. Conclusion The established LAMP technique in this study enables rapid,sensitive and specific detection of Nm,LAMP-based detection,where amplification was completed within 60 min,faster than PCRS. Therefore,LAMP does not require expensive device,nor does it require skilled workers,which is suitable for the application in grass-root and small-scale laboratories as well as field surveillance.
作者
闫永飞
黄亮
赵丽萍
刘晓娟
丁国涛
胡朝辉
YAN Yongfei;HUANG Liang;ZHAO Liping;LIU Xiaojuan;DING Guotao;HU Zhaohui(Handan Center for Disease Control and Prevention,Hebei 056008,China)
出处
《医学动物防制》
2018年第11期1087-1089,1092,共4页
Journal of Medical Pest Control
基金
邯郸市科学技术局立项-邯郸市流脑菌带菌研究及快速检测方法的建立(1623208066-6)
关键词
环介导等温扩增
脑膜炎奈瑟菌
ctrA基因
Loop-mediated isothermal amplification
Neisseria meningitids
ctrA gene
