摘要
为建立一种用于快速检测鸡传染性法氏囊病病毒(IBDV)的逆转录环介导等温扩增技术(RTLAMP)方法,设计了针对VP1基因保守序列的5组引物,采用荧光检测试剂通过实时浊度仪监测阳性扩增结果。结果显示,从5组引物中筛选出了1组对IBDV有效扩增的引物,利用该引物能够在63℃、1 h条件下实现IBDV VP1基因片段的特异性扩增,与其他病毒的核酸无扩增反应;样品RNA的最低检出量为5×10-5ng/L;利用建立的检测方法对14份临床样品进行检测,其阳性检出率为100%。结果表明,建立的IBDV RT-LAMP检测方法具有快速、准确、特异性强、灵敏度高和便捷的特点,可用于IBD临床样品的检测。
To establish a reverse transcription loop-mediated isothermal amplification(RT-LAMP)technique for rapid detection of infectious bursal disease virus(IBDV),five groups of primers,including two primer sets,targeting the conserved sequence of VP1 gene,were designed.The positive amplification results were monitored by a real-time turbidimeter using the fluorescence detection reagent.In result,one of the group of primers for efficient amplification of IBDV was screened out from the five primers groups,and the primers were specific in amplification of the IBDV VP1 gene fragment within 1 h at 63 ℃.The primers showed no amplification reaction with nucleic acids of other viruses.And the minimum detectable amount of sample RNA was 5×10^-5ng/L.Finaly,fourteen IBD clinical samples were tested using the established assay and the positive detection rate was 100%.The results showed that the established IBDV RT-LAMP detection method is a fast,accurate,specific,sensitive and convenient technique,which can be a potential method for the detection IBDV in clinical IBD samples.
作者
陈艳艳
陈盛峰
陈果
陈博文
何秀苗
韦平
CHEN Yan-yan;CHEN Sheng-feng;CHEN Guo;CHEN Bo-wen;HE Xiu-miao;WEI Ping(Guangxi Colleges and Universities Key Laboratory of Utilization of Microbial and Botanical Resources ,School of Marine Sciences and Biotechnology ,Guangxi University for Nationalities ,Nanning 530006 ,China;Guangxi Key Laboratory Cultivation Base for Polysaccharide Materials and Modifications,Guangxi University for Nationalities ,Nanning 530006, China;Institute for Poultry Science and Health,Guangxi University,Nanning 530005,China)
出处
《中国兽医科学》
CAS
CSCD
北大核心
2019年第2期169-175,共7页
Chinese Veterinary Science
基金
国家自然科学基金项目(31560706
31660717)
广西科技重大专项(桂科AA17204057)
广西自然科学基金项目(2017GXNSFAA198033
2014GXNSFDA118018)
2014广西民族大学相思湖青年学者创新团队项目(2014)
广西民族大学2018年研究生教育创新计划项目(gxun-chxzs2018066)
关键词
传染性法氏囊病病毒
RT—LAMP
实时浊度仪
检测
infectious bursal disease virus
reverse transcription loop-mediated isothermal amplification
real-time turbidimeter
detection
