重组酶聚合酶扩增技术快速检测鲤春病毒血症病毒(SVCV)方法的建立  

Recombinase polymerase amplification for the rapid detection of Spring viremia of carp virus(SVCV)

作　　者：梁君妮 尹伟力 谢爽 刘宁 段效辉 林森 LIANG Jun-ni;YIN Wei-li;XIE Shuang;LIU Ning;DUAN Xiao-hui;LIN Sen(Technology Center of Yantai Customs,Yantai 264000,China)

机构地区：[1]烟台海关技术中心

出　　处：《中国预防兽医学报》2019年第10期1037-1040,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家重点研发计划(2017YFF0211103);国家质量监督检验检疫总局科研项目(2017IK232)

摘　　要：为建立鲤春病毒血症病毒(SVCV)的快速检测方法,本研究根据SVCV的G基因保守序列设计特异性引物,通过条件优化初步建立了基于重组酶聚合酶扩增技术(RPA)的SVCV检测方法。该RPA方法最优扩增温度为30℃,恒温反应20 min内即可完成。特异性试验结果显示,该方法与其它常见鱼类感染病毒无交叉反应;敏感性试验结果显示,该方法对SVCV质粒标准品的最低检出限为89.2拷贝/μL,高于传统的RT-PCR方法;利用已建立的RPA方法和RT-PCR方法对市场购买的80份临床样品进行检测,结果显示,RPA方法与RT-PCR方法检测结果一致,且RPA方法能够有效检出RT-PCR方法的弱阳性样品,表明RPA方法可以用于临床检测。本研究建立的检测SVCV的RPA方法具有快速、简便、特异性强、灵敏度高的特点,适合基层实验室及现地检测。In order to develop a rapid method for the detection of spring Viremia of carp virus(SVCV), the specific primers were designed by targeting the G gene of SVCV. The recombinase polymerase amplification(RPA) was established by optimizing the reaction conditions. The amplification temperature of the RPA method was determined at 30℃, and the test can be finished within 20 minutes. This method is specific with no cross-reaction with other common fish infection viruses. The detection limit of this method was 89.2 copies/μL, which was higher than that of the traditional RT-PCR method. Moreover, a total of 80 clinical samples were detected by both the RPA method and the RT-PCR method, respectively. The weak positive samples tested by RT-PCR method could be detectable with the RPA method, it is indicated that the RPA method could be used in clinical detection.This method is rapid, simple, specific and sensitive for testing SVCV, and it will be widely used in the Grassroots laboratory and On-site inspection.

关 键 词：重组酶聚合酶扩增技术 快速检测 鲤春病毒血症病毒 

分 类 号：S852.65[农业科学—基础兽医学]