甲型流感病毒荧光RT-PCR检测方法的设计和检定  

Design and rapid evaluation of a TaqMan assay for the detection of influenza A viruses

作　　者：陈继明[1] 王志亮[1] 逄增昌[2] 孙承英[1] 孙映雪[1] 宋翠平[1] 弋英[1] 

机构地区：[1]农业部动物检疫所国家外来动物疫病诊断中心,青岛266032 [2]青岛市疾病控制中心

出　　处：《中华实验和临床病毒学杂志》2005年第1期80-83,共4页Chinese Journal of Experimental and Clinical Virology

基　　金：国家十五重大攻关项目资助 (2 0 0 2BA5 14A 18) ;青岛市重大应急项目资助 (0 3 ny 2 0 )

摘　　要：目的　设计甲型流感病毒TaqMan荧光RT-PCR检测方法并对其进行检定。方法　用DNAStar和PrimerPremier5. 0软件设计甲型流感病毒荧光PCR检测所用引物和探针 ;在GenBank中进行Blast以及电子对比证明其具有高度的特异性和保守性 ;和标准RT-PCR进行比较 ,检测此方法的灵敏度。结果　所设计的引物和探针高度特异和保守 ,灵敏度比标准RT-PCR方法高 3～ 27倍 ,并且反转录和PCR可合并为一步。结论　设计了甲型流感病毒TaqMan检测方法 ;该方法具有特异。Objective To design and rapidly evaluate a TaqMan assay for detecting influenza A viruses. Methods The probe and the primers of the assay were designed with the software packages of DNA Star and Primer Premier 5.0. Their specificity and conservation were verified through Blast in GenBank and electronic hybridization. The assay′s sensitivity was compared with the standard RT-PCR. Results The designed primers and probe were confirmed to be very specific and conserved. The assay was 3-27 folds more sensitive than the standard RT-PCR. The RT and PCR steps could be simplified into one step. Conclusion The TaqMan Real-time PCR assay is specific, sensitive and easy to perform.

关 键 词：甲型流感病毒 荧光PCR RT-PCR方法 引物 特异性 探针 灵敏 荧光RT-PCR检测 高度 反转录 

分 类 号：S852.65[农业科学—基础兽医学]