摘要
基于A型和C型鸭甲肝病毒(Duck hepatitis A virus,DHAV)3D基因的6个保守区域设计了4条引物,利用BstDNA聚合酶在60℃恒温保持45min即可完成反转录和扩增反应,由此建立了鸭甲肝病毒的一步反转录环介导等温扩增(RT-LAMP)方法。该方法具有良好的特异性,除DHAV外,对其他4种常见鸭病的检测结果均为阴性。该方法对DHAV-A和DHAV-C病毒RNA的最低检出量均为0.1pg,是常规RT-PCR方法的10倍。临床应用结果表明,该方法与病毒分离鉴定方法的符合率为92.5%,而且对仪器要求低,适于基层实验室和现场检测。
An one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for detecting duck hepatitis A virus (DHAV) was established with four primers based on six conserved positions of the 3D gene.The process of assay was completed by using Bst DNA within 45 min at 60℃.The detect limit of the RT-LAMP assay was 0.1 pg of viral RNA,which was 10 times higher than RT-PCR,and no amplification was found with the samples of 4 other common duck diseases.The clinical application results showed the coincidence rate between the assay and the virus isolation and identification is 92.5%.The assay was a potential useful technique for DHAV detection in field condition.
作者
扈琴吉
杨宏禹
路晓
于可响
孙晓军
HU Qinji;YANG hongyu;LU Xiao;YU Kexiang;SUN Xiaojun(Institute of Poultry,Shandong Academy of Agricultural Sciences,Jinan 250023,China;Daliuhang Veterinary Station of Shandong Province,Weihai 265600,China)
出处
《家禽科学》
2019年第6期38-42,共5页
Poultry Science
基金
国家重点研发计划(2016YFD0500800)
国家重点研发计划(2016YFD0500100)
山东省现代农业产业技术体系家禽创新团队计划(SDAIT-11-01)
山东省农业科学院创新工程项目(CXGC2016B14)
关键词
鸭甲肝病毒
3D基因
一步反转录环介导等温扩增
仪器要求低
Duck hepatitis A virus
3D gene
One-step reverse transcription loop-mediated isothermal amplification
Low requirement for instrument
