生物学杂志

生物学杂志 ›› 2025, Vol. 42 ›› Issue (6): 93-.doi: 10.3969/j.issn.2095-1736.2025.06.093

• 技术方法 • 上一篇    下一篇

基于RPA分析辅助CRISPR/Cas12a系统的WSSV检测方法的建立与应用

牟全玲， 丁存宝   

  1. 华北理工大学 生命科学学院， 唐山 063210
  • 出版日期:2025-12-18 发布日期:2025-12-19
  • 通讯作者: 丁存宝，博士，副教授，研究方向为斑马鱼毒理病理学，E-mail:10332282@qq.com
  • 作者简介:牟全玲，硕士研究生，研究方向为对虾病毒检测，E-mail:1455530172@qq.com
  • 基金资助:
    唐山市重点研发计划项目（22150206A）

Establishment and application of WSSV detection method based on RPA analysis assisted CRISPR/Cas12a system

MU Quanling, DING Cunbao   

  1. School of Life Sciences, North China University of Science and Technology, Tangshan 063210, China
  • Online:2025-12-18 Published:2025-12-19

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摘要： 研究建立一种针对白斑综合征病毒（white spot syndrome virus, WSSV）的基于重组酶聚合酶等温扩增（recombinant polymerase amplification, RPA）结合CRISPR/Cas12a系统及侧流层析试纸条（lateral flow strip, LFS）的检测方法，即RPA-CRISPR/Cas12a-LFS，该方法具有快速、灵敏、操作简单等特点。RPA-CRISPR/Cas12a-LFS 的检测方法下限是每个反应1.12×101copies的白斑综合征病毒DNA，并且整个过程可以在1 h内完成。此外，这种检测组分与其他常见对虾病毒和健康对虾基因组均没有交叉反应。从对虾养殖场采集的110个临床对虾样本，检测结果与实时聚合酶链式反应测试结果完全一致。研究构建的检测方法（RPA-CRISPR/Cas12a-LFS）作为对虾中WSSV早期诊断的新型检测方案具有巨大潜力。

关键词: 白斑综合征病毒, 重组酶聚合酶等温扩增, CRISPR-Cas12a系统, 侧流层析试纸条, 病毒检测

Abstract: n this study, an RPA-CRISPR/Cas12a-LFS assay was developed for the detection of white spot syndrome virus （WSSV） based on recombinase polymerase amplification （RPA） combined with CRISPR/Cas12a system and lateral flow strips （LFS）, which was rapid, sensitive and simple to operate. The lower limit of the detection method for RPA-CRISPR/Cas12a-LFS was 1.12×101copies of vitiligo syndrome virus DNA per reaction, and the whole process could be completed within 1 h. Furthermore, the assay showed no cross-reactivity with other common shrimp viruses or healthy shrimp genomes. When testing 110 clinical shrimp samples collected from shrimp farms, the results of this study were completely consistent with the results of real-time polymerase chain reaction （PCR） assays. In conclusion, the detection method （RPA-CRISPR/Cas12a-LFS） constructed in this study shows great potential as a novel detection scheme for the early diagnosis of WSSV in shrimp.

Key words: white spot syndrome virus, recombinant polymerase amplification, CRISPR/Cas12a, lateral flow strip, virus detection

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