口蹄疫病毒通用型RT-RAA-LFD快速检测方法的建立
Development of a Universal RT-RAA-LFD Method for Rapid Detection of FMDV
寇美玲 1谢佳芮 1李占鸿 1张振兴 1苏晓航 1王轶男 1宋建领 1苗海生1
作者信息
- 1. 云南省畜牧兽医科学院,云南省热带亚热带动物病毒病重点实验室,云南昆明 650224
- 折叠
摘要
为建立一种操作简便、有效的口蹄疫病毒(FMDV)核酸通用型检测方法,以FMDV 3D基因为靶基因,设计标记引物及探针,建立了一种新的检测FMDV的反转录-重组酶介导链置换核酸扩增结合侧流层析试纸检测(RT-RAA-LFD)方法.该方法操作简便,耗时短,40℃35 min内即可完成检测,通过肉眼观察可判定结果;具有较好的覆盖面,能够检出我国O型、A型FMDV主要流行拓扑型病毒,与猪瘟病毒(CSFV)、猪伪狂犬病病毒(PRV)等其他猪病病原无交叉反应;对RNA标准品的最低检测限为 10-7 拷贝/μL,敏感性较高.利用建立的FMDV RT-RAA-LFD方法对 5 份临床样品进行检测,其结果与RT-PCR结果一致.结果表明,本研究建立的FMDV RT-RAA-LFD方法具有快速、直观、特异强、重复性好等优点,适合在基层兽医防疫部门和养殖场推广应用.
Abstract
In order to develop a universal method to rapidly and effectively detect the foot and mouth disease virus(FMDV),a new method of reverse transcription recombinase-aided amplification combined with lateral flow dipstick(RT-RAA-LFD)was developed by designing labeling primers and probes targeting at FMDV 3D gene.The established method was efficient and convenient,which could complete detection within 35 min at 40℃and determine results through visual observation;with good coverage,major prevalent topological viruses including FMDV O and A type in China could be detected,and no cross-reaction with other swine pathogens such as classical swine fever virus(CSFV)and porcine pseudorabies virus(PRV)was observed;and the lowest detection limit to RNA standard substances was 10-7 copies/μL,with high sensitivity.5 clinical samples were detected by the established FMDV RT-RAA-LFD method,receiving consistent results with those by RT-PCR.In conclusion,the established method,with the advantages of rapidness,visualization,strong specificity and good repeatability,was appropriate for grass-roots animal disease prevention departments and farms.
关键词
口蹄疫病毒/重组酶介导链置换核酸扩增/侧流层析试纸/恒温扩增/快速检测Key words
FMDV/RAA/LFD/isothermal amplification/rapid detection引用本文复制引用
基金项目
国家重点研发计划项目(2021YFD1800300)
云南省科技人才与平台计划项目(202205AF150007)
出版年
2024
国家科技期刊平台
NSTL
万方数据