摘要：根据G en Bank中已登陆的猪流行性腹泻病毒(PED V)、猪传染性胃肠炎病毒(TGEV)和猪轮状病毒(RV)的参考基因序列,设计3对引物分别用于扩增PEDV的N基因、TGEV的M基因、RV的VP7基因的目的片段;通过优化反应体系,建立了同时检测PEDV、TGEV、RV的三重PCR检测方法。敏感性和特异性试验的结果表明,该方法对这3种病毒的最低核酸检出量分别为39.5 pg(PEDV)、38.1 pg(TGEV)和51.6 pg(RV)。该方法可同时扩增477 bp(PEDV)、263 bp(TGEV)、355 bp(RV)的特异性基因片段,而对猪瘟病毒(CSF V)、猪繁殖与呼吸综合征病毒(PRRSV)、猪圆环病毒2型(PCV2)和伪狂犬病病毒(PRV)等病毒的检测结果均为阴性。三重PCR与单一PCR的符合率为100%。上述结果表明,该方法的建立对于这3种病毒的临床快速检测与流行病学调查具有十分重要的意义。

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。

摘要：Objective] This study aimed to develop a reverse transcription loop-medi-ated isothermal amplification （RT-LAMP） method for detecting BVDV. [Method] Since gp48 gene of BVDV is among the most conserved regions, a set of four primers was designed to amplify six target sequences at the gp48 gene region for the RT-LAMP assay. The optimization of the RT-LAMP reaction was performed by evaluat-ing reaction temperature and reaction time. [Result] The RT-LAMP aasay was suc-cessful y conducted at 56 ℃ within 40 min under isothermal conditions, and the re-sults could be detected as ladder-like bands using agarose gel electrophoresis. The RT-LAMP assay is highly sensitive and able to detect 3.74 ×100 copies/μl of BVDV RNA, as no cross-reaction was observed with other viruses. [Conclusion] Overal , the newly established RT-LAMP assay indicates the potential application in both clinical diagnosis and field surveil ance of BVDV.