猪流行性乙型脑炎病毒纳米PCR检测方法的建立  

Establishment of nano PCR assay for detection of the Japanese encephalitis virus

作　　者：柴瑞影 华荣虹[2,3] 袁万哲[1,2,4] 孙继国[1,2,4] 

机构地区：[1]河北农业大学动物医学院,河北保定071001 [2]河北省兽医生物技术工程技术研究中心,河北保定071001 [3]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室,黑龙江哈尔滨150001 [4]农业部动物疫病病原生物学华北区观测实验站,河北保定071001

出　　处：《中国兽医学报》2017年第3期471-474,共4页Chinese Journal of Veterinary Science

基　　金：兽医生物技术国家重点实验室开放基金资助项目(SKLVBF201407)

摘　　要：根据猪流行性乙型脑炎病毒(JEV)E基因的保守区域设计1对扩增270bp片段的特异性检测引物,将纳米金颗粒添加到PCR反应体系中,同时优化反应条件,建立了高效、灵敏的纳米PCR检测方法。采用该方法灵敏度可达3.26×10~2拷贝/μL,普通PCR方法仅为3.26×10~4拷贝/μL,且与其他病毒没有交叉感染。试验结果表明,该纳米PCR检测方法不仅特异性好,而且其敏感性是普通PCR检测方法的100倍。采用纳米PCR方法和普通PCR方法对临床送检的32份样品进行检测,检测结果纳米PCR检出2份阳性样品,普通PCR检测均为阴性。A rapid and sensitive nanoparticle-assisted polymerase chain reaction （nanoPCR） assay for detection of the Japanese encephalitis virus was established（JEV）. According to the conserved region of the E gene,one set of specific primers was designed to amplify a 270 bp fragment,the sensitivity and specificity of the method were investigated. Using the optimized conditions for de- tection of JEV RNA, the lowest detection limit of the nano PCR assay was about 3.26）〈 102 copies/μL and the conventional PCR assay was about 3.26 ）〈 104 copies/μL, and no cross-reaction was ob- served with other viruses. The sensitivity test showed that the sensitivity of nano-PCR was more than 100 times than that of conventional PCR. A total of 32 clinical samples were detected by this assay,and the results showed that two samples were positive by the nano PCR and all the samples were negative by the conventional PCR.

关 键 词：猪流行性乙型脑炎病毒 纳米PCR E基因 

分 类 号：S852.65[农业科学—基础兽医学]