H6亚型禽流感病毒荧光定量RT-PCR检测方法的建立和应用

Development and application of one step fluorescent quantitative RT-PCR for H6 subtype avian influenza virus

为建立一种快速检测H6亚型禽流感病毒的基因检测方法,根据Gen Ban K中H6亚型禽流感病毒的HA基因保守序列设计一对引物和一条Taq Man探针,通过各项条件优化建立了检测H6亚型禽流感病毒的实时荧光定量RT-PCR方法。结果显示,该方法特异性强,与其它亚型(H1、H3、H4、H5、H7、H9)禽流感病毒、传染性法氏囊病毒、鸡传染性支气管炎病毒、鸡新城疫病毒、马立克氏病毒、传染性喉气管炎病毒等均不发生交叉反应。该方法经实验室和临床上多次试验,证明其特异性强、重复性好、灵敏度高,可用于H6亚型流感病毒的监测。

In order to establish a gene detection method for rapid detection of H6 subtype avian influenzavirus,a pair of primers and a Taq Man probe were designed based on the conservative sequence of the HA gene of H6 subtype avian influenza virus in GenBanK. The real-time fluorescent quantitative RT-PCR method for detecting H6 subtype avian influenza virus was established through various conditions. The re-sults showed that the method had a strong specificity and no cross reaction with other subtypes （H1,H3,H4,H5,H7,H9） avian influenza virus,infectious bursal virus,infectious bronchitis virus,Newcastle dis-ease virus,Marek＇s virus,infectious laryngotracheitis virus and so on. The method has been proved to be highly specific,reproducible and highly sensitive by laboratory and clinical trials. It can be used for moni-toring H6 subtype influenza virus.