Shandong Provincial Key Laboratory of Biochemical Engineering, College of Marine Science and Biological Engineering, Qingdao University of Science and Technology, Qingdao, 266042, PR China.
College of Life Sciences, Qingdao University, Qingdao, 266071, PR China.
Talanta. 2020 Aug 15;216:120978. doi: 10.1016/j.talanta.2020.120978. Epub 2020 Mar 31.
Amplification reactions coupled with electrophoresis or real-time fluorescence system have been successfully used for the analysis of nucleic acid. However, complex procedures or expensive instruments greatly limit their application in low-resource settings. To address these shortcomings, we fabricated a universal and simple detection platform by integrating strand exchange amplification (SEA) with lateral flow assay (LFA) strip. SEA is a simple isothermal amplification reaction, only requires a pair of primers and one DNA polymerase, above all, its short amplicons are easy to migrate on the strip. LFA strip was proved to be stable for months without large signal deviations and result could be easy to read by naked eyes, which makes it an appropriate option for field-based analysis. Our proposed SEA-LFA strip could reliably detect as few as 0.05 nM pork DNA and 0.07 nM duck DNA by the naked eye, its analytical performance is comparable with laboratory-based real-time fluorescence test. Furthermore, this proof-of-concept method could be applied to detect a wide variety of nucleic acid by focusing on primer design rather than on the development of a wholly new analysis platform. We believe that this simple visualization system has great potential as a preliminary test tool in resource-limited areas.
扩增反应与电泳或实时荧光系统相结合已成功用于核酸分析。然而，复杂的程序或昂贵的仪器极大地限制了它们在资源有限环境中的应用。为了解决这些缺点，我们通过将链交换扩增（SEA）与横向流动分析（LFA）条带相结合，制造了一种通用且简单的检测平台。SEA 是一种简单的等温扩增反应，仅需要一对引物和一种 DNA 聚合酶，最重要的是，其短的扩增子很容易在条带上迁移。LFA 条带已被证明可以稳定数月而不会出现大的信号偏差，并且结果可以通过肉眼轻松读取，这使其成为现场分析的合适选择。我们提出的 SEA-LFA 条带可以通过肉眼可靠地检测低至 0.05 nM 的猪 DNA 和 0.07 nM 的鸭 DNA，其分析性能可与基于实验室的实时荧光测试相媲美。此外，这种概念验证方法可以通过专注于引物设计而不是开发全新的分析平台来应用于检测各种核酸。我们相信，这种简单的可视化系统具有作为资源有限地区初步测试工具的巨大潜力。
