鸭瘟病毒环介导等温扩增(LAMP)快速检测方法的建立与应用  

Development of a loop-mediated isothermal amplification assay for the rapid detection of duck plague virus

作　　者：张坤[1] 刁有祥[1] 鞠小军[1] 

机构地区：[1]山东农业大学动物科技学院,山东泰安271018

出　　处：《中国兽医学报》2013年第4期520-525,共6页Chinese Journal of Veterinary Science

基　　金：现代农业产业技术体系专项资金资助(CARS-43-34)

摘　　要：为建立一种简便、快速、灵敏、特异的鸭瘟病毒(DPV)检测方法,本研究根据GenBank中登录的DPV基因组序列,设计3对特异性的环介导等温扩增(LAMP)引物,经优化反应体系,建立了LAMP快速检测方法。结果表明,LAMP方法能够在63℃恒温下,1h内实现目的核酸的大量扩增。结果判定时只需要在扩增产物中加入SYBRGreenⅠ染料,就可以直接在可见光或紫外光下观察颜色变化。该方法敏感性可达0.245μg/L,比普通PCR灵敏性高10倍;对鸭病毒性肝炎病毒、H9N2亚型禽流感病毒、鸭副黏病毒、鸭源偏肺病毒等的核酸无交叉反应;利用建立的LAMP检测方法对40份临床样品的阳性检出率为30%。该方法简便快捷,省时省力,是一种适用于基层实验室快速、准确检测DPV的方法。To develop a rapid and sensitive method for detecting duck plague virus（DPV）,a loopmediated isothermal amplification （LAMP） assay was established using three pairs of specific primers. The assay was optimized to amplify DPV DNA by incubation at 63℃ for lh. The results visualized directly or under the UV light with added SYBR Green I dye. The diagnostic method was sensitive and specific,the amplification results of duck hepatitis virus, H9N2 avian influenza virus,duck paramyxovirus and duck avian metapneumovirus were negative,and its lowest detection limit was 0. 245/2g/L,which was 10-fold higher than the conventional PCR. The positive rate of forty duck samples suspected duck plague was 30%. Therefore the LAMP assay developed in this study provided a rapid and practical method for DPV detection.

关 键 词：鸭瘟病毒(DPV) 环介导等温扩增技术(LAMP) 

分 类 号：S852.657[农业科学—基础兽医学]