食品与发酵工业 >

2022 , Vol. 48 >Issue 17: 291 - 297

DOI: https://doi.org/10.13995/j.cnki.11-1802/ts.031606

分析与检测

重组酶聚合酶扩增反应结合侧流层析试纸条技术快速鉴别大西洋鳕鱼制品真伪

  • 王慧芳 ,
  • 喻勇新 ,
  • 李晨虹 ,
  • 陈文隽 ,
  • 凌岚馨 ,
  • 周颖
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  • 1(上海海洋大学 食品学院,上海,201306)
    2(农业农村部水产品贮藏保鲜质量安全风险评估实验室,上海,201306)
    3(上海海洋大学 环境DNA技术与水生态健康评估工程中心,上海,201306)
    4(上海海洋大学 海洋动物系统分类与进化上海高校重点实验室,上海,201306)
第一作者:本科生(李晨虹教授为通信作者,E-mail:chli@shou.edu.cn)

收稿日期: 2022-03-18

  修回日期: 2022-04-24

  网络出版日期: 2022-10-01

基金资助

上海市科学技术委员会2022年度“创新行动计划”农业领域项目(22N31900700);上海市科学技术委员会部分地方院校能力建设项目(19050501900)

收起

Real-time characterization of Atlantic cod by recombinase polymerase amplification coupled lateral flow dipstick (RPA-LFD)

  • WANG Huifang ,
  • YU Yongxin ,
  • LI Chenhong ,
  • CHEN Wenjun ,
  • LING Lanxin ,
  • ZHOU Ying
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  • 1(College of Food Science and Technology, Shanghai Ocean University, Shanghai 201306, China)
    2(Laboratory of Quality and Safety Risk Assessment for Aquatic Products on Storage and Preservation, Ministry of Agriculture and Rural Affairs, Shanghai 201306, China)
    3(Engineering Research Center of Environmental DNA and Ecological Water Health Assessment, Shanghai Ocean University, Shanghai 201306, China)
    4(Shanghai Universities Key Laboratory of Marine Animal Taxonomy and Evolution, Shanghai Ocean University, Shanghai 201306, China)

Received date: 2022-03-18

  Revised date: 2022-04-24

  Online published: 2022-10-01

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摘要

建立了重组酶聚合酶扩增反应(recombinase polymerase amplification,RPA)结合侧流层析试纸条技术(RPA-lateral flow dipstick,RPA-LFD)快速鉴别大西洋鳕鱼制品真伪的方法。对比大西洋鳕鱼和常用来冒充鳕鱼的鱼类线粒体基因组序列,设计针对大西洋鳕鱼的特异性探针和引物,建立针对大西洋鳕鱼的RPA-LFD检测方法。该实验筛选最佳引物和探针并评估其特异性,优化反应温度、时间和现场检测条件,并对20份市售“大西洋鳕鱼”及常见的冒充鳕鱼的鱼类制品进行现场检测,评估检测效果。实验结果表明,RPA-LFD检测方法特异性强,在44 ℃、14 min条件下的检测效果最好。在现场开展快速检测,效果良好,20份鳕鱼制品检测结果经过基因测序证实和产品真实成分一致。该研究建立的大西洋鳕鱼RPA-LFD检测方法特异性强、反应耗时短、操作简便、结果可视,有良好的现场快速检测应用前景,为大西洋鳕鱼制品现场快速检测提供了新思路,为水产品市场监管提供了技术支持。

关键词： 大西洋鳕鱼; 重组酶聚合酶扩增反应; 真伪; 侧流层析试纸条技术; 现场快速检测

本文引用格式

王慧芳 , 喻勇新 , 李晨虹 , 陈文隽 , 凌岚馨 , 周颖 . 重组酶聚合酶扩增反应结合侧流层析试纸条技术快速鉴别大西洋鳕鱼制品真伪[J]. 食品与发酵工业, 2022 , 48(17) : 291 -297 . DOI: 10.13995/j.cnki.11-1802/ts.031606

Abstract

In this study, a method of recombinase polymerase amplification (RPA) combined with lateral flow dipstick (LFD) technology (RPA-LFD) was established for the characterization of Atlantic cod related products. Based on the comparison between the sequences of mitochondrial genomes of the Atlantic cod and popular adulterated cod on the market, specific probes and primers were designed for detecting the Atlantic cod and an RPA-LFD detection method for the Atlantic cod were established. The objectives of this study were to select the best primers and probes, evaluate their specificity and optimize the best reaction conditions, which including temperature, reaction time both in the lab and in the field. The optimized method was used to perform the real-time detection of 20 ‘cod' products and common adulterated ones that were currently available on the market. The results demonstrated that RPA-LFD showed a strong specificity by incubating the reaction at 44 ℃ for 14 min. The results of real-time detection of 20 ‘cod' products were consistent with the true identity of the product, which was confirmed by sequencing results. The method of RPA-LFD for distinguishing Atlantic cod and related products established in this study showed a strong specificity and with the advantages of short reaction time, simple protocol, and visible results. It provides a new strategy for real-time identification of cod products in the field, which is important for the supervision of the market of aquatic products.

Key words： Gadus morhua; recombinase polymerase amplification; authenticity; lateral flow dipstick; real-time detection in the field

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