An Innovative and Efficient Fluorescent Detection Technique for Salmonella in Animal-Derived Foods Using the CRISPR/Cas12a-HCR System Combined with PCR/RAA

文献类型： 外文期刊

第一作者： Wang, Yuanshang

作者： Wang, Yuanshang;Shao, Yanchun;Wang, Xiaohong;Wang, Yuanshang;Shao, Yanchun;Wang, Xiaohong;Wang, Yuanshang;Du, Pengfei;Wang, Weiting;Liu, Yaobo;Ma, Yanli;Hu, Peng;Cao, Jianfang;Abd El-Aty, A. M.;Abd El-Aty, A. M.

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关键词： CRISPR/Cas12a-HCR; PCR/RAA; specific target; Salmonella; fluorescence rapid detection; animal-derived foods

期刊名称：JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY （ 影响因子：6.1； 五年影响因子：6.3 ）

ISSN： 0021-8561

年卷期： 2024 年 72 卷 15 期

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收录情况： SCI

摘要： Here, we present a method for Salmonella detection using clustered regularly interspaced short palindromic repeats associated with the CRISPR-associated protein 12a-hybridization chain reaction (CRISPR/Cas12a-HCR) system combined with polymerase chain reaction/recombinase-assisted amplification (PCR/RAA) technology. The approach relies on the Salmonella invA gene as a biorecognition element and its amplification through PCR and RAA. In the presence of the target gene, Cas12a, guided by crRNA, recognizes and cleaves the amplification product, initiating the HCR. Fluorescently labeled single-stranded DNA (ssDNA) H1 and H2 were introduced, and the Salmonella concentration was determined based on the fluorescence intensity from the triggered HCR. Both assays demonstrate high specificity, sensitivity, simplicity, and rapidity. The detection range was 2 x 10(1)-2 x 10(9) CFU/mL, with an LOD of 20 CFU/mL, and the entire process enabled specific and rapid Salmonella detection within 85-105 min. Field-incurred spiked recovery tests were conducted in mutton and beef samples using both assays, demonstrating satisfactory recovery and accuracy in animal-derived foods. By combining CRISPR/Cas12a with hybridization chain reaction technology, this study presents a rapid and sensitive Salmonella detection method that is crucial for identifying pathogenic bacteria and monitoring food safety.

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