应用TaqMan-BHQ探针实时荧光RT-PCR法定量检测马秋波病毒核酸

Application of TaqMan-BHQ Probe on Quantitative Detection for Machupo Virus by Real-time Fluorescence RT-PCR

〔目的〕建立一种适应口岸马秋波病毒实时荧光定量RT-PCR快速的检测方法。〔方法〕用专业软件设计引物和TaqMan-BHQ探针,以人工合成马秋波病毒S基因的片段作为模板,进行实时荧光定量RT-PCR研究。〔结果〕模板的Ct值与模板稀释浓度的对数存在良好的线性关系,标准曲线为Y=-3.281X+50.975,R2=0.999361,PCR扩增效率为101.0%,其最低检出限为28 copies/μl。〔结论〕应用TaqMan-BHQ1探针的实时荧光RT-PCR检测马秋波病毒核酸,具有耗时短、灵敏度高等特点。

Objective To establish a rapid and accurate method of real-time fluorescence RT-PCR to quantify the S gene of Machupo virus.Methods The twin primers and the TaqMan-BHQ probe were designed and synthesized based on the S gene segment of Machpo virus.The real-time RT-PCR was developed for detecting machupo virus.Results The Ct value of templates had a linear relationship with the log starting quantity.Sensitivity assay showed that the detection limit of the assay was 28 copies/μl and the standard curve Y=-3.281X＋50.975 had a good reproducibility.Conclusions Quantitative real-time fluorescence for Machupo virus detection was established and characterized by rapidity and sensitivity.