鸡传染性法氏囊病病毒实时荧光定量RT-PCR检测方法的建立

Development of a real-time fluorescent quantitative RT-PCR for detection of infectious bursal disease virus

本试验根据GenBank中鸡传染性法氏囊病病毒(IBDV)基因组设计一对针对病毒vp5基因的引物和一条特异性Taq Man探针,建立了一种快速检测IBDV核酸载量的Taq Man荧光定量RT-PCR方法。通过对反应条件和反应体系的优化,使得该方法在108拷贝/μL～101拷贝/μL范围内具有良好的线性关系。能够灵敏地检测初始模板中30个拷贝的病毒核酸,其灵敏度是常规RT-PCR检测方法的100倍。该方法不与其它的禽源病毒发生非特异性反应,并且具有良好的重复性,为IBDV的定性定量检测提供了有效的工具。

A real-time fluorescent quantitative RT-PCR was developed for the detection of infectious bursal disease virus （IBV） using a pair of primers and a specific Taq Man probe specific to the vp5 gene of IBV. The assay was optimized to produce linearity from 10^8 copies/μL to 10^1 copies/μL in standard curve. It is highly specific and no cross reactivity was detected against other virus. It had a sensitivity of detecting 30 copies of viral RNA. The assay developed here could provide a powerful tool for quantification of IBDV.