蜜蜂黑色王台病毒SYBR GreenⅠRT-qPCR检测方法的建立

A Real-time Quantitative RT- PCR Assay for the Detection of Black Queen-Cell Virus in Honeybee

为建立蜜蜂黑色王台病毒RT-qPCR检测方法,以蜜蜂黑色王台病毒(BQCV)衣壳蛋白基因部分序列作为目的基因,构建标准质粒并作为模板,建立标准曲线,且对所建立方法的灵敏度和特异性进行分析。结果表明:所建立的RT-qPCR检测方法,扩增效率E=1 08%,可信度R2=0.997。该方法灵敏度高,最低检出量为101个拷贝;该方法特异性强,仅能检出BQCV,而不能检测出蜜蜂囊状幼虫病毒、畸翅病毒。以上结果表明已成功建立了针对BQCV灵敏、特异的RT-qPCR检测方法。

To establish a bee black queen cell virus fluorescence real-time quantitative RT-PCR detection methods, by building honey black queen cell virus （BQCV） the capsid protein gene partial sequence of standard plasmid as a template for the establishment of a standard curve, and established the specificity and sensitivity of the method was tested. The results showed that： fluorescence quantitative RT-PCR amplification efficiency of the established E=108% , R2=0.997, the minimum detectable amount of 10^1 copies; this method can only be detected BQCV virus, but can not detect cystic bee larvae virus, abnormal wing virus, with good specificity. Show has successfully established a specific BQCV sensitive real-time quantitative RT-PCR detection method.