Abstract：
Capripoxvirus (CaPV) contains three viruses that have caused massive losses in the livestock and dairy industries. Accurate CaPV differentiation has far-reaching implications for effectively controlling outbreaks. However, it has a great challenge to distinguishing three viruses due to high homology of 97%. Here, we established a sensitive CRISPR/Cas12a array based on Multiple-recombinase polymerase amplification (M-RPA) for CaPV differentiation, which provided a more comprehensive and accurate differentiation mode targeting VARV B22R and RPO30 genes. By sensitive CRISPR/Cas12a and M-RPA, the actual detection limits of three viruses were as low as 50, 40 and 60 copies, respectively. Moreover, Lateral flow dipstick (LFD) array based on CRISPR/Cas12a achieved portable and intuitive detection, making it suitable for point-of-care testing. Therefore, CRISPR/Cas12a array and LFD array paved the way for CaPV differentiation in practice. Additionally, we constructed a real-time quantitative PCR (qPCR) array to fill the qPCR technical gap in differentiation and to facilitate the quarantine departments.
摘要：
Capripoxvirus（CaPV）包含三种病毒，它们在畜牧业和乳制品行业造成了巨大损失。准确的CaPV分化对有效控制疫情具有深远的意义。然而,由于具有97％的高同源性，因此区分三种病毒具有很大的挑战。这里,我们建立了一个敏感的CRISPR/Cas12a阵列基于多重重组酶聚合酶扩增（M-RPA）的CaPV分化，针对VARVB22R和RPO30基因提供了更全面、准确的分化模式。通过敏感的CRISPR/Cas12a和M-RPA，三种病毒的实际检测限低至50、40和60个拷贝，分别。此外，基于CRISPR/Cas12a的侧流试纸(LFD)阵列实现了便携直观的检测,使其适合点的护理测试。因此,CRISPR/Cas12a阵列和LFD阵列在实践中为CaPV分化铺平了道路。此外,我们构建了实时定量PCR（qPCR）阵列，以填补qPCR在区分和方便检疫部门的技术空白。
