南方医科大学学报
南方醫科大學學報
남방의과대학학보
JOURNAL OF SOUTHERN MEDICAL UNIVERSITY
2009年
8期
1545-1547
,共3页
莫秋华%杨翠兰%林继灿%谭华%涂承宁%叶立青%刘志明%杜坚%孙虹%杨泽
莫鞦華%楊翠蘭%林繼燦%譚華%塗承寧%葉立青%劉誌明%杜堅%孫虹%楊澤
막추화%양취란%림계찬%담화%도승저%협립청%류지명%두견%손홍%양택
多重RT-PCR%A型流感病毒%B型流感病毒%甲型H1N1流感病毒
多重RT-PCR%A型流感病毒%B型流感病毒%甲型H1N1流感病毒
다중RT-PCR%A형류감병독%B형류감병독%갑형H1N1류감병독
multiplex RT-PCR%influenza%type A influenza virus%type B influenza virus%novel A (H1N1) influenza virus
目的 建立能同时筛查A型、B型和新型甲型H1N1流感病毒的多重RT-PCR技术.方法 针对A型流感病毒的M基因、B型流感病毒的NS基因设计通用扩增引物,针对新型甲型H1N1流感病毒的HA基因设计特异性扩增引物,采用一步法建立多重RT-PCR反应体系.通过盲法实验与实时荧光RT-PCR进行比对来评价方法的准确性,并应用于临床评价方法的实用性和有效性.结果 琼脂糖凝胶电泳分析多重RT-PCR产物,结果显示目的扩增片段条带清晰明亮,没有非特异性产物出现,可见该方法扩增效率高,特异性强.50份样本的盲法实验结果显示两种方法检测结果完全一致,符合率为100%.结论 建立的多重RT-PCR方法能通过一次实验快速、准确地同时筛检A型、B型和新型甲型H1N1流感病毒,是一项成本低廉、对流感的疫情监测和早期诊断具有实用价值的筛检技术.
目的 建立能同時篩查A型、B型和新型甲型H1N1流感病毒的多重RT-PCR技術.方法 針對A型流感病毒的M基因、B型流感病毒的NS基因設計通用擴增引物,針對新型甲型H1N1流感病毒的HA基因設計特異性擴增引物,採用一步法建立多重RT-PCR反應體繫.通過盲法實驗與實時熒光RT-PCR進行比對來評價方法的準確性,併應用于臨床評價方法的實用性和有效性.結果 瓊脂糖凝膠電泳分析多重RT-PCR產物,結果顯示目的擴增片段條帶清晰明亮,沒有非特異性產物齣現,可見該方法擴增效率高,特異性彊.50份樣本的盲法實驗結果顯示兩種方法檢測結果完全一緻,符閤率為100%.結論 建立的多重RT-PCR方法能通過一次實驗快速、準確地同時篩檢A型、B型和新型甲型H1N1流感病毒,是一項成本低廉、對流感的疫情鑑測和早期診斷具有實用價值的篩檢技術.
목적 건립능동시사사A형、B형화신형갑형H1N1류감병독적다중RT-PCR기술.방법 침대A형류감병독적M기인、B형류감병독적NS기인설계통용확증인물,침대신형갑형H1N1류감병독적HA기인설계특이성확증인물,채용일보법건립다중RT-PCR반응체계.통과맹법실험여실시형광RT-PCR진행비대래평개방법적준학성,병응용우림상평개방법적실용성화유효성.결과 경지당응효전영분석다중RT-PCR산물,결과현시목적확증편단조대청석명량,몰유비특이성산물출현,가견해방법확증효솔고,특이성강.50빈양본적맹법실험결과현시량충방법검측결과완전일치,부합솔위100%.결론 건립적다중RT-PCR방법능통과일차실험쾌속、준학지동시사검A형、B형화신형갑형H1N1류감병독,시일항성본저렴、대류감적역정감측화조기진단구유실용개치적사검기술.
Objective To developed a multiplex RT-PCR assay for simultaneous screening of type A, B and novel A (H1N1)influenza viruses. Methods Two pairs of universal primers in were designed for amplifying the M gene and NS gene of type A and B influenza viruses, respectively. A pair of specific primers of HA gene was designed to detect novel A (H1N1) influenza virus. A one-step method was used to establish the multiplex RT-PCR system. A blinded experiment was carried out to validate the accuracy of this assay in comparison with the results of real-time fluorescence RT-PCR. The clinical practicability and efficacy of this assay was also evaluated. Results The RT-PCR products were analyzed using agarose gel electrophoresis,which yielded distinct bands of the target fragments without non-specific reactions, suggesting the high efficiency and specificity of the multiplex RT-PCR. Blinded study of 50 samples demonstrated a concordance rate of 100%. Conclusion This multiplex RT-PCR assay allows one-step simultaneous detection of type A, B and novel A (H1N1) influenza viruses rapidly and accurately, and provides a valuable low-cost screening technique for influenza epidemic monitoring and early diagnosis.
