郑州大学学报（理学版）2026，Vol.58Issue(1)：87-94,8.DOI:10.13705/j.issn.1671-6841.2024197
基于RPA-CRISPR/Cas12a的猴痘病毒基因检测方法建立
Establishment of a RPA-CRISPR/Cas12a-based Gene Detection Method for Monkeypox Virus
摘要
Abstract
A rapid monkeypox virus nucleic acid detection system was developed by integrating recombi-nase polymerase isothermal amplification(RPA)and clustered regularly interspaced short palindromic re-peats-associated protein 12a(CRISPR/Cas12a).The system was combined with lateral flow assay(LFA)to construct an RPA-CRISPR/Cas12a-based rapid nucleic acid detection platform for MPXV,en-abling visual and rapid detection.The optimal reaction conditions for RPA were determined to be 37℃for 15 minutes.For the CRISPR system,the optimal volumes of Cas12a,crRNA,NEBuffer,and ssDNA were identified as 1.0,0.25,3.0,and 0.25 μL,respectively.The entire detection process was comple-ted within 1 hour,and no cross-reactivity was observed with inactivated influenza A/B virus stock solu-tions or plasmid controls for vaccinia and variola.The limit of detection(LOD)was confirmed to be 10 copies/μL.A highly sensitive,specific,and rapid RPA-CRISPR/Cas12a-based method for MPXV de-tection was successfully developed.The method was demonstrated to be applicable to on-site testing and was proven a simple and rapid solution for MPXV detection.By integrating isothermal amplification,CRISPR-based specificity,and LFA visualization,the practicality of this method in resource-limited set-tings was significantly enhanced.The study highlights the potential of this approach as a promising tool for early outbreak control and public health surveillance.关键词
猴痘病毒/簇状规则间隔短回文重复序列相关蛋白12a/重组酶聚合酶等温扩增技术/胶体金试纸条Key words
MPXV/CRISPR/Cas12a/RPA/colloidal gold test strip分类
农业科技引用本文复制引用
董小旭,牛楠楠,赵印震,杜学利,李灵轲,王继创,李玉林,王云龙..基于RPA-CRISPR/Cas12a的猴痘病毒基因检测方法建立[J].郑州大学学报（理学版）,2026,58(1):87-94,8.基金项目
河南省科技攻关项目(232102311036) （232102311036）
