摘要
[目的 ]快速灵敏检测猪伪狂犬病毒(PRV)[方法 ]根据PRV g B基因设计特异性引物,建立了PRV纳米PCR检测方法。[结果 ]PRV纳米PCR方法的最低检出限为10拷贝/μL,其敏感性比未添加纳米金的对照PCR提高100倍;建立的纳米PCR与猪细小病毒、猪圆环病毒Ⅱ型、猪繁殖与呼吸综合征病毒等常见猪病毒性病原均无交叉反应。应用纳米PCR和PRV国家标准中的PCR方法,对2014—2016年期间采集自17个省市发病猪场的148份临床样品进行平行检测,PRV纳米PCR的阳性检出率为49.3%(73/148),PRV国标PCR方法的阳性检出率为23.6%(35/148),并且PRV纳米PCR检测为阳性,而PRV国标PCR方法检测为阴性的38份样品,测序结果均确认为PRV阳性。[结论 ]本研究建立的纳米PCR检测方法敏感特异,可用于猪伪狂犬病的病原学检测。
[Objective]In order to develop a rapid and sensitive method for detecting Pseudorabies virus（PRV）.[Methods]A nano PCR assay for detection of PRV was developed with a pair of specifi c primers that were designed based on sequence of g B gene.[Results]Results showed the detection limit of this assay was 10 copies/μL,sensitivity of which was 100 times higher than conventional PCR assay that no gold nanoparticle was added. Good specifi city was also identifi ed that there was no cross reaction with other common viral pathogens of swine such as porcine parvovirus（PPV）,porcine circovirus type 2（PCV2）,porcine reproductive and respiratory syndrome virus（PRRSV）. Using PRV nano PCR and national standard PCR test of g B gene,148 clinical samples collected from 17 different areas during 2014-2016 were conducted parallel detection. The positive rates by nano PCR and national standard PRV were 43.5%（73/148）and 27.2%（35/148）. 38 samples tested negative by national standard PRV presented a positive result when using the method of nano PCR,which were consistent with DNA sequencing.[Conclusion]The established PRV nano PCR method was applicable for pathogenic detection of PRV due to its high sensitivity and good specifi city.
出处
《中国动物检疫》
CAS
2017年第3期102-105,共4页
China Animal Health Inspection
基金
科技部国家重点研发计划(2016YFD0501501)
