猴痘病毒PCR检测方法的建立

从GenBank上调取猴痘病毒的基因序列,经过分析,找出了猴痘病毒的特异性靶基因序列F3L片段,人工合成猴痘病毒MPV(AF380138)F3L基因片段(48048-48509bp)并插入质粒,作为病毒检测的模拟阳性模板。根据该序列设计并合成PCR引物,对模拟的阳性模板进行扩增,结果能扩增出与目的片段大小一致的条带。建立了PCR检测方法,经条件优化后,对鸡痘、禽痘、羊痘等14种相似病毒核酸进行PCR扩增,发现具有良好的特异性。PCR方法能扩增出0.3pg的阳性模板,显示建立的PCR方法具有高效、快速、特异、灵敏的特点,可用于口岸猴痘病毒的检疫。

Establishment of PCR Assays for the Detection of Monkeypox Virus

The F3L gene sequence ofmonkeypox virus was synthesized and cloned into the plasmid to be used as a target. A pair of PCR primers were designed and synthesized based on the nucleotide sequences of MPV in GenBank. After optimization of the detection methods, they were used to amplify MPV. The results showed that the primers could get the specific bands. The high specificity of the methods was demonstrated by detecting 14 viruses, such as chickenpox, goalpox and so on. The maximal detection limits by the primers were approximately 0.3 pg of plasmid DNA. Therefore, PCR method developed here for MPV detection had the characteristic of high efficiency, speediness, specificity and sensitivity. They can be used to entry-exit quarantine detection ofmonkeypox virus.