应用多重PCR技术快速鉴定登革病毒

Rapid detection of dengue viruses by multiplex PCR assay

目的建立一套应用多重PCR技术快速分型鉴定4种血清型登革病毒的方法.方法 用随机引物将1～4型登革病毒RNA逆转录成cDNA,再将4对登革病毒型特异性引物同时加到一个反应管中进行多重PCR扩增,根据扩增片段的大小判断血清型,并用2004年收集的登革热病人血清标本验证新建立的方法.结果 同时用4对登革病毒型特异引物进行多重PCR,1～4型登革病毒标准毒株相应扩增的片段大小分别为391、319、216和152 bp,与预期的扩增片段大小相符.2004年4份登革热病人血清标本扩增的片段大小均为391 bp,属1型登革病毒.结论 新建立的方法是一种特异、快速的登革病毒分型鉴定方法.

Objective To establish a multiplex PCR assay for rapid detection and typing of 4 serotypes of dengue virus. Methods Random primer was used to reverse transcribe the 4 types of dengue viral RNA, then a multiplex PCR assay was proceeded to amplify the cDNA by adding 4 pairs of dengue virus type - specific primers into one reaction tube. Based on the length of the PCR amplicon, all 4 dengue virus serotypes can be identified. To evaluate the new established method, serum samples of dengue fever patients collected in 2004 were tested. Results The lengths of the PCR amplicon of dengue virus type 1 to 4 were 391 bp, 319 bp, 216 bp and 152 bp respectively. The length of amplicon in all samples collected in2004 was 391 bp, which in dicated the serotype of dengue virus in that year was type 1. Conclusion The established method is a specific and rapid detection and typing assay for dengue virus.