中国兽医科学2024，Vol.54Issue(6)：735-741,7.DOI:10.16656/j.issn.1673-4696.2024.0088
基于RPA-CRISPR/Cas12a的异尖线虫快速可视化检测方法
RPA-CRISPR/Cas12a-based visual and rapid detection of Anisakis
摘要
Abstract
To achieve rapid,simple,and efficient detection of Anisakis in seafood to control its prevalence,recombinase polymerase amplification(RPA)primers and CRISPR RNA(crRNA)for the conserved sequence of internal transcribed space in Anisakis were designed,the RPA-CRISPR/Cas12a fluorescence detection system was established and reaction conditions such as crRNA concentration were optimized.The detection limit,specificity,and repeatability of this method were evaluated,and its actual detec-tion ability was verified through artificially contaminated samples.The results showed that the RPA-CRISPR/Cas12a fluorescence detection system could complete the reaction within 40 min,and with the sensitivity of 1 copy/μL.This method could only detect Anisakis and not react with Hysterothy-lacium aduncum,Ligula intestinalis,Trichiurus lepturus tissue and Larimichthys polyactis tissue.The RPA-CRISPR/Cas12a fluorescence visualization detection system was fast,sensitive,and specific,and could be applied to the detection of Anisakis in marine products.关键词
异尖线虫/RPA/CRISPR/Cas12a/可视化检测Key words
Anisakis/RPA/CRISPR/Cas12a/fluorescent detection分类
农业科技引用本文复制引用
王皓璐,赵连静,陈秀琴,杨亚明,吴方伟,刘晓雷,王洋,白雪..基于RPA-CRISPR/Cas12a的异尖线虫快速可视化检测方法[J].中国兽医科学,2024,54(6):735-741,7.基金项目
"十三五"国家重点研发计划项目(2017YFC1601206) （2017YFC1601206）
云南省科技人才与平台计划(院士专家工作站)项目(202305AF150167) （院士专家工作站）
