Division of Swine Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, China.
J Virol Methods. 2013 Nov;193(2):374-8. doi: 10.1016/j.jviromet.2013.07.018. Epub 2013 Jul 18.
Nanoparticle-assisted polymerase chain reaction (nanoPCR) is a novel method for the rapid amplification of DNA and has been adopted for the detection of virus because of its simplicity, rapidity, and specificity. A nanoPCR assay was developed to detect and differentiate wild-type and gene-deleted pseudorabies virus (PRV). Three pairs of primers for nanoPCR developed in this study were selected from conserved regions of PRV, producing specific amplicons of 431 bp (gB), 316 bp (gE), and 202 bp (gG). The sensitivity of this assay using purified plasmid constructs containing the specific gene fragments was 100-1000 fold higher than conventional PCR. The PRV nanoPCR assay did not amplify porcine parvovirus, porcine circovirus type 2, porcine reproductive and respiratory syndrome virus, porcine teschovirus, or African swine fever virus but produced three bands of expected size with PRV and two bands of expected size with the gene-deleted PRV-Bartha-K61. Of 110 clinical samples collected from seven provinces in China, 53% and 48% were positive for wild-type PRV according to the nanoPCR assay and virus isolation, respectively.
纳米颗粒辅助聚合酶链反应（nanoPCR）是一种快速扩增 DNA 的新方法，因其简单、快速和特异性而被用于病毒检测。本研究开发了一种 nanoPCR 检测方法，用于检测和区分野毒株和基因缺失型伪狂犬病病毒（PRV）。本研究设计了 3 对用于 nanoPCR 的引物，针对 PRV 的保守区域，产生了 431bp（gB）、316bp（gE）和 202bp（gG）的特异性扩增子。使用含有特定基因片段的纯化质粒构建体进行的该检测方法的灵敏度比常规 PCR 高 100-1000 倍。该 PRV nanoPCR 检测方法不能扩增猪细小病毒、猪圆环病毒 2 型、猪繁殖与呼吸综合征病毒、猪传染性胃肠炎病毒或非洲猪瘟病毒，但与 PRV 和基因缺失型 PRV-Bartha-K61 均能产生预期大小的 3 条和 2 条条带。在中国七个省份采集的 110 个临床样本中，nanoPCR 检测和病毒分离的野毒株阳性率分别为 53%和 48%。
