甲型H1N1流感病毒FAST-LAMP检测方法的建立

摘要

建立一种新型等温扩增检测方法,用于提高等温扩增反应的检测速度,应用于甲型H1N1流感病毒检测.根据HA基因设计一组引物,包括外引物、内引物、环引物及套内引物,套内引物的加入增加了引物与模板的接触位点,提高扩增效率,缩短检测时间.结果显示,FAST-LAMP检测法比普通LAMP检测法时间缩短45 min左右,比加入环引物的LAMP检测方法缩短20 min,大大缩短了反应的检测时间.检测灵敏度可达到1×102拷贝.FAST-LAMP是一种高效、更快的检测方法.%It was to establish a new loop-mediated isothermal amplification(FAST-LAMP)for rapidly detecting influenza A(H1N1) virus. A set of primers were designed according to HA gene, including the outer primers, internal primers, loop primers and nest internal primers. The nest internal primers increased the contact sites of primers and nucleic acid template, so that the amplification efficiency could be improved and detection time reduced. The FAST-LAMP method could be 45 less than LAMP method, and 20 minutes less than LAMP method with loop primers, i.e. detection time reduced significantly detecting sensitivity reached 1×102 copies cDNA template/tube. It proved that the established FAST-LAMP method is an efficient and reliable method for detecting influenza A(H1N1)virus.