施马伦贝格病毒环介导等温扩增快速检测方法的建立

Development of rapid assay for Schmallenberg virus by loop-mediated isothermal amplification method

目的 建立施马伦贝格病毒逆转录环介导等温扩增（RT-LAMP）快速检测方法.方法 从GenBank数据库中获得施马伦贝格病毒S基因序列,应用DNAStar软件MegAlign程序分析其序列,利用PrimerExplorerV4软件在序列保守区域设计LAMP引物,即外引物、内引物,建立施马伦贝格病毒等温扩增快速检测方法.结果 RT-LAMP检测方法对施马伦贝格病毒RNA的灵敏度达到10个拷贝,所用引物对赤羽病毒、口蹄疫病毒、蓝舌病毒、水泡性口炎病毒及牛病毒性腹泻病毒无特异性扩增,表现出良好特异性.结论 建立的施马伦贝格病毒等温扩增快速检测方法灵敏度高、特异性强,为快速检测施马伦贝格病毒提供了新方法.

Objective To set up loop-mediated isothermal amplification（RT-LAMP） method for quick detection of Schmallenbergvirus.Methods The S gene sequences of Schmallenbergvirus were downloaded from GenBank database and analyzed using MegAlign tools of software DNAStar.The outer primers and inner primers of LAMP were then designed according to their conservative region using software PrimerExplorer V4,and the reaction conditions were optimized.Results The results showed that the RTLAMP assay had a detection limit of 10 RNA copies.The primers used in this assay showed the specificity for Schmallenberg virus and did not amplify FMDV,BVDV,BTV,AKAV and VSV.Conclusion It is indicated the RT-LAMP method is a simple,rapid,accurate,sensitive and specific method for detecting Schmallenberg virus.This assay offers a new method and idea for detecting Schmallenberg virus.