用于诊断4种猪病毒病的寡核苷酸芯片的研制

制备可同时检测引起集约化养猪业常见4种传染病,病毒(猪流感病毒、口蹄疫病毒、猪伪狂犬病毒、猪蓝耳)的寡核苷酸芯片。根据4种猪疫病病毒特异性基因的保守区域设计合成了60 mer寡核苷酸探针,制备寡核苷酸芯片。采用不对称PCR和间接荧光标记技术进行单链DNA扩增和荧光标记,标记样品与寡核苷酸芯片杂交后,进行芯片清洗、扫描及结果分析。杂交结果显示,4种病毒的寡核苷酸检测探针均特异地与相应的标记样品杂交,芯片上呈现较强的阳性杂交信号,而除阳性质控探针外,阴性对照和空白对照均检测不到荧光信号。证明寡核苷酸芯片适用于快速、准确、高通量地诊断影响集约化养猪业的多种猪疫病病毒。

Development of oligonucleotide microarray for detection of SIV, FMDV, PRV and PRRSV

An oligonucleotide microarray was developed to detect simultaneously four severe epidemic porcine viruses,including swine influenza virus(SIV),foot-and-mouth disease virus(FMDV),porcine pseudorabies virus(PRV) and porcine reproductive and respiratory syndrome virus(PRRSV).The 60-mer oligonucleotide probes based on specific conservative DNA fragments of the four porcine viruses were designed and synthesized,then applied to fabricating the oligonucleotide microarrays.By means of asymmetric PCR and indirect fluorescent-labeling and technology to amplify and label the samples,they were hybridized with the oligonucleotide microarrays,followed by washing,scanning and analysis.The hybridization results showed the detected probes of the four porcine viruses could specifically hybridize with the corresponding sample with strong hybridization signals,while negative and blank controls could not detect any fluorescent signals.The results suggest the 60mer oligonucleotide microarrays can be applied to rapidly,exactly and high-throughput detecting several infectious porcine viruses.