跨越式滚环扩增(SRCA)结合金纳米粒子可视化检测食品中的沙门氏菌
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(1.河北农业大学食品科技学院 河北保定 071001;2.河北农业大学理工系 河北沧州 061100;3.河北农业大学生命科学学院 河北保定 071001;4.河北省人畜共患病原微生物分析与防控重点实验室 河北保定 071000)

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国家自然科学基金项目(32172288,31371772);河北省自然科学基金重点项目(C2019204342);中央引导地方科技发展资金项目(216Z5501G);河北省外专百人计划项目(360-0803-JSN-3YGS)


Visual Detection of Salmonella in Food Using Saltatory Rolling Circle Amplification Combined with Gold Nanoparticles
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(1.School of Food Science and Technology, Hebei Agricultural University, Baoding 071001, Hebei;2.Department of Science and Engineering, Hebei Agricultural University, Cangzhou 061100, Hebei ;3.College of Life Sciences, Hebei Agricultural University, Baoding 071001, Hebei ;4.Key Laboratory of Microbiological Analysis and Control of Human and Livestock Communities in Hebei Province,Baoding 071000, Hebei)

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    摘要:

    沙门氏菌是引起全球人类腹泻病的主要病因,严重危害人畜健康。传统检测方法耗时长,步骤繁琐,因此开展快速简便灵敏的可视化检测方法,具有重要意义。本文设计能与靶序列结合的探针,将其与金纳米粒子(AuNPs)结合。经跨越式滚环等温扩增技术(SRCA)扩增的靶序列,高温变性后成为单链,其与金纳米粒子上的探针结合后,释放出金纳米粒子,在MgSO4的作用下,聚集呈现肉眼可见的蓝色,判定为阳性,阴性为紫色。依此,建立一种简便、快捷、灵敏的可视化检测方法。结果表明:该方法特异性良好,能够区分8株沙门氏菌阳性与8株非沙门氏菌阴性。可视化检测的灵敏度为5.5×100 CFU/mL。将牛奶样品人工污染沙门氏菌,检出限为3.7×100 CFU/mL。在50份实际样品检测中,与GB 4789.4-2016方法进行比较,该方法敏感性为100.00%,特异性为97.87%,符合率达到98.00%。本文建立的SRCA-AuNPs可视化检测方法,具有较高的应用价值,结果肉眼可见,适合于现场可视化检测和基层单位使用。

    Abstract:

    Salmonella is the main cause of human diarrhea worldwide, which seriously endangers human and animal health. Traditional detection methods are time-consuming and cumbersome, so it is of great significance to carry out rapid, simple and sensitive visual detection methods. In this paper, a probe that can bind to the target sequence was designed and combined with gold nanoparticles (AuNPs). The target sequence was amplified by saltatory rolling circle amplification (SRCA). After denaturation at high temperature, it became a single chain. After binding with the probe on gold nanoparticles, gold nanoparticles were released. Under the action of MgSO4, the aggregation showed blue visible to the naked eye, which was judged to be positive and negative to be purple. Therefore, a simple, fast and sensitive visual detection method was established. The results showed that the method had good specificity and could distinguish 8 Salmonella positive and 8 non-Salmonella negative. The sensitivity of visual detection was 5.5×100 CFU/mL. The milk samples were artificially contaminated with Salmonella, and the detection limit was 3.7×100 CFU/mL. Comparison with GB 4789.4-2016 method for detection of 50 actual samples, the sensitivity of this method was 100.00%, the specificity was 97.87%, and the coincidence rate was 98.00%. The SRCA-AuNPs visual detection method established in this paper has high application value, and the results are visible visually, which is suitable for on-site visual detection and grassroots units.

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董贺楠,张蕴哲,徐慧,檀建新,张伟,袁耀武.跨越式滚环扩增(SRCA)结合金纳米粒子可视化检测食品中的沙门氏菌[J].中国食品学报,2024,24(3):240-248

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  • 收稿日期:2023-03-25
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  • 在线发布日期: 2024-04-16
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