牛疱疹病毒Ⅰ型(BHV-1)荧光定量PCR检测方法的建立及应用  

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

作　　者：王吉[1] 付瑞[1] 李晓波[1] 王淑菁[1] 王莎莎[1] 李威[1] 秦骁 黄宗文 巩薇[1] 岳秉飞[1] 贺争鸣[1] WANG Ji;FU Rui;LI Xiaobo;WANG Shujing;WANG Shasha;LI Wei;QIN Xiao;HUANG Zongwen;GONG Wei;YUE Bingfei;HE Zhengming(National Institutes for food and drug Control,National Center for quality of Laboratory Animal,Beijing 100050,China)

机构地区：[1]中国食品药品检定研究院国家实验动物微生物遗传检测中心

出　　处：《实验动物科学》2019年第5期35-41,46,共8页Laboratory Animal Science

基　　金：中国食品药品检定研究院学科带头人基金项目(No.2015X5)

摘　　要：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 g B基因设计特异引物和Taq Man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。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 g B 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 × 10^1 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.

关 键 词：牛疱疹病毒Ⅰ型 实时荧光定量PCR 牛源性样本 

分 类 号：R372[医药卫生—病原生物学]