猪繁殖与呼吸综合征病毒与猪流行性腹泻病毒双重RT-PCR检测方法的建立与应用

发布时间：2018-03-19 16:19

本文选题：猪繁殖与呼吸综合征病毒　切入点：猪流行性腹泻病毒　出处：《中国兽医科学》2017年05期 　论文类型：期刊论文

【摘要】：为快速简便地检测猪繁殖与呼吸综合征病毒(PRRSV)与猪流行性腹泻病毒(PEDV),根据GenBank中已发布的基因序列,选择PRRSV的ORF7基因和PEDV的S基因,分别设计1对特异性引物,通过对反应体系中各成分的优化,建立了可同时检测PRRSV和PEDV的双重RT-PCR方法。本方法能从PRRSV和PEDV的混合基因组中分别扩增出300 bp和578 bp的特异性片段,且对猪传染性胃肠炎病毒、猪瘟病毒、乙型脑炎病毒、轮状病毒、大肠杆菌的扩增结果均为阴性;应用该方法对采自成都周边的79份临床样品进行检测,结果与单一RT-PCR的检测结果一致,具有良好的重复性;本方法检出的最小拷贝数为1.49×10~5copies/μL,敏感性也较好。本研究建立的双重RT-PCR方法具有较好实用性,可用于临床样品的检测。
[Abstract]:In order to detect porcine reproductive and respiratory syndrome virus (PRRSVV) and porcine epidemic diarrhea virus (PEDVV) quickly and easily, according to the published gene sequence in GenBank, the ORF7 gene of PRRSV and the S gene of PEDV were selected and a pair of specific primers were designed. By optimizing the components of the reaction system, a dual RT-PCR method was established for simultaneous detection of PRRSV and PEDV. The specific fragments of 300bp and 578bp were amplified from the mixed genome of PRRSV and PEDV, respectively. The amplification results of classical swine fever virus, Japanese encephalitis virus, rotavirus and Escherichia coli were all negative. This method has good reproducibility, the minimum copy number of this method is 1.49 脳 10 ~ (5) copi es / 渭 L, and the sensitivity is good. The dual RT-PCR method established in this study has good practicability and can be used for the detection of clinical samples.
【作者单位】：四川农业大学动物医学院;四川农业大学动物疫病与人类健康四川省重点实验室;
【基金】：四川省科技支撑计划项目(2017NZ0038) “十二五”农村领域国家科技计划项目(2015BAD12B04-2.3)
【分类号】：S852.651

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