犬Ⅰ型腺病毒荧光定量PCR检测方法的建立

Establishment of Real-time Fluorescent Quantitative PCR Assay for Detection of Canine Adenovirus TypeⅠ

通过分析对比GenBank公布的几株犬Ⅰ型腺病毒序列,针对犬Ⅰ型腺病毒特有的E3区,设计了1对特异性引物。对引物浓度、退火温度、扩增体系等条件优化后,建立了犬Ⅰ型腺病毒荧光定量PCR检测方法,并对40份CAV-Ⅰ人工感染银黑狐的粪便样品进行了检测。结果表明,与普通PCR相比,建立的犬Ⅰ型腺病毒荧光定量PCR检测方法具有特异性好、灵敏度高、重复性强的特点。该检测方法可用于Ⅰ型犬腺病毒的快速诊断与监测,并能够定量分析CAV-Ⅰ的感染程度。

In order to establish a real-time fluorescent quantitative PCR assay for rapid detection of canine adenovirus typeⅠ(CAV-Ⅰ),the special primers were designed for E3 conserved region based on the sequences of several strains of canine adenovirus typeⅠin GenBank,and the concentration of primer,Tm value and volume of amplification were optimized.We used this established assay to detect the CAV-Ⅰin forty fecal samples from artificially infected silver foxes.Compared to the general PCR assay,results showed that the real-time fluorescent quantitative PCR assay for CAV-Ⅰis more specific,highly sensitive and repeatable.Therefore,it could be applied for rapid detection,monitor and quantitative analysis of canine adenovirus typeⅠ.