牛疱疹病毒Ⅰ型(BHV-1)荧光定量PCR检测方法的建立及应用

目的 建立牛疱疹病毒Ⅰ型(BHV-1)实时荧光定量PCR检测方法，用于牛源性样本中BHV-1的快速检测。方法 根据已发表的BHV-1 gB基因设计特异引物和TaqMan探针，建立BHV-1实时荧光定量PCR方法。并对方法的特异性、敏感性、重复性稳定性等进行测定。用建立的方法对181份牛源性样本进行检测。结果 建立的BHV-1荧光定量PCR检测方法与牛副流感病毒Ⅲ型(BPIV3)、牛病毒性腹泻病毒1型(BVDV1)、猪伪狂犬病毒(PRV)、单纯疱疹病毒Ⅰ型(HSV-1)、猫疱疹病毒1型(FHV-1)均无交叉反应；检测灵敏度可达到1×101copies/μL；批内变异系数均小于5%。应用建立的方法检测181份牛源性样本，有6份样本BHV-1核酸为阳性。结论 建立的BHV-1荧光定量PCR检测方法具有快速、特异、敏感及稳定的特点，可用于牛源性样本中BHV-1污染的检测。

Establishment and Application of Real-time Fluorescent Quantitative PCR Method for Detection of Bovine Herpesvirus Type 1

Objective To establish a real-time fluorescent quantitative PCR method for detection of Bovine herpesvirus type 1 (BHV-1)in Bovine origin samples.Method The primers and TaqMan probe were designed and synthesized according to the published BHV-1 specific sequences of gB gene.Q-PCR method is established.Then carries on the specificity,sensitivity,repeatability and stability of this method were tested.The method is used to detect 181 Bovine origin samples.Result The developed Q-PCR method was no cross reaction with Bovine parainfluenza virus type 3 (BPIV3),Bovine viral diarrhea virus type 1(BVDV1),herpes virus type 1 (HSV-1),Felid herpesvirus 1 (FHV-1),and Pig pseudo rabies virus (PRV);sensitivity was 1×101 copies/μL;Coefficient of variation(CV)was less than 5%;There were 6 positive reaction detected in the 181 Bovine origin samples.Conclusion The developed PCR method is good in specificity,sensitivity,repeatability and stability and may be used for rapid quantitative detection the BHV-1 in Bovine origin samples.