Highly efficient gene editing of Feline herpesvirus 1 using CRISPR/Cas9 combined with FACS

文献类型： 外文期刊

第一作者： Wang, Hai-Ming

作者： Wang, Hai-Ming;Cai, Bing-Yan;Lu, Hui;Xu, Shi-Jia;Tang, Yan-Dong;Qiu, Wen-Ying

作者机构：

关键词： Feline herpesvirus 1 (FHV-1); CRISPR/Cas9; fluorescence-activated cell sorting (FACS); homologous recombination; gene knockout

期刊名称：FRONTIERS IN CELLULAR AND INFECTION MICROBIOLOGY （ 影响因子：4.8； 五年影响因子：5.5 ）

ISSN： 2235-2988

年卷期： 2025 年 15 卷

页码：

收录情况： SCI

摘要： Feline herpesvirus 1 (FHV-1) is a major causative agent of feline viral rhinotracheitis and ocular lesions. Due to its large DNA genome, the construction of recombinant FHV-1 viruses presents considerable challenges for conventional methodologies. In this study, we implemented an integrated strategy combining CRISPR/Cas9-mediated gene editing with fluorescence-activated cell sorting (FACS) to enable the rapid and efficient generation of recombinant FHV-1 viruses. Specifically, the thymidine kinase (tk) gene was disrupted by inserting a monomeric Cherry (mCherry) reporter gene, and the glycoprotein E (gE) gene was similarly interrupted through the incorporation of a green fluorescent protein (GFP) reporter. The CRISPR/Cas9 system enables precise, site-specific genomic modifications, while FACS allows for effective enrichment and isolation of the desired recombinant viral populations. This combined approach significantly reduces the time required for recombinant virus generation from weeks to days, thereby offering substantial potential to expedite vaccine development and advance functional genomics research.

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