农业生物技术学报2026，Vol.34Issue(1)：199-211,13.DOI:10.3969/j.issn.1674-7968.2026.01.017
基于PCR-LFS的CRISPR/Cas9基因编辑元件快速检测技术及其在转基因作物监测中的应用
PCR-LFS-Based Rapid Detection Technology for CRISPR/Cas9 Gene-editing Elements and Its Application in Transgenic Crop Monitoring
摘要
Abstract
The widespread application of CRISPR/Cas9 gene editing technology in crop breeding has demonstrated significant potential for improving crop resistance,yield,and quality.However,this progress has also heightened the need for effective detection of gene-edited components.A significant number of crop samples containing non-excised exogenous CRISPR/Cas9 elements remain prevalent during early breeding stages and under specific conditions.However,existing detection methods are associated with limitations such as low sensitivity,time-consuming procedures,or dependence on sophisticated instrumentation.To meet the growing demand for rapid detection of CRISPR/Cas9-edited crops,a highly sensitive method integrating PCR with lateral flow strips(PCR-LFS)was developed in this study.Cas9-specific primers were designed and the amplification system was optimized,allowing rapid visual detection using colloidal gold-labeled test strips.A detection limit of 0.012％(by mass fraction)and sensitivity of 17.5 copies/μL were achieved,outperforming conventional PCR-capillary gel electrophoresis(0.1％)and quantitative PCR(1.75×104 copies/μL).Specificity testing confirmed that Cas9-positive tomato(Solanum lycopersicum)samples were accurately distinguished from non-transgenic crops without cross-reactivity.Practical validation demonstrated that reliable detection was achieved in mixed samples at a ratio of 1∶213,with high repeatability and stability.Compared with traditional techniques,this method eliminated the need for complex instruments,reduced detection time to 3 h,and was suitable for on-site screening and port inspections.It was suggested that future optimization of primer design and LFS procedures might extend the application to other gene-edited components.This study provides strong technical support for precise monitoring of genetically modified crops.关键词
CRISPR/Cas9基因编辑作物/PCR与侧向层析流试纸条(PCR-LFS)/灵敏度/特异性/快速可视化检测Key words
CRISPR/Cas9-gene edited crops/PCR with lateral flow strips(PCR-LFS)/Sensitivity/Specificity/Rapid visualization assay分类
农业科技引用本文复制引用
高婕妤,韩芳,曾德新,余晓峰,丁柳,孔维恒,尤征合,李云飞,宗凯,孙娟娟,余华峥,胡康棣..基于PCR-LFS的CRISPR/Cas9基因编辑元件快速检测技术及其在转基因作物监测中的应用[J].农业生物技术学报,2026,34(1):199-211,13.基金项目
国家重点研发计划(2023YFF0611500) （2023YFF0611500）
安徽省重点研究与开发计划(2022i01020001) （2022i01020001）
