改良多重实时荧光定量PCR检测流感A型和B型病毒

摘要

Objective To set up a modified multiplex real-time fluorescence quantitative PCR ( MRT-PCR) assay to detect influenza virus A and B rapidly .Methods The MRT-PCR assay was established according to target sequence of influenza virus A and B specific gene selected by bioinformatics analysis results .The standard products and quality control samples were built to evaluate the sensitivity , specificity and repeatability of the system .Results The MRT-PCR detection system was successfully established based on Taqman-molecular beacon ( Taqman-MB) probe and Homo-Taq Assidted Non-Dimer ( HAND) system.The detection limit of the assay was 102 copies/μl,which showed good specificity and repeatability , with coefficient of variation (CV) being 0.99%~2.50%.Conclusion A fast,specific,sensitive,stable, modified MRT-PCR detection system is successfully established ,which possesses favourable application prospect in clinical practice .%目的：建立改良多重实时荧光定量PCR体系快速检测流感A型和B型病毒。方法根据生物信息学分析结果选定流感A型和B型病毒的靶序列，建立MRT－PCR检测体系。构建质粒标准品，评估所建立体系的灵敏度、特异性和重复性。结果成功建立了基于同源加尾系统和Taqman－分子灯标探针的改良多重实时荧光定量PCR检测体系；该方法最低检测限为102 copies／μl，特异性良好，重复性良好，变异系数（CV）为0．99％～2．50％。结论建立了快速、敏感、特异、稳定地改良多重实时荧光定量PCR（ MRT－PCR）检测体系，具有良好的临床应用前景。