College of Plant Protection, Nanjing Agricultural University, Nanjing 210095, China.
College of Forestry, Nanjing Forestry University, Nanjing 210037, China.
Plant Dis. 2020 Nov;104(11):2774-2778. doi: 10.1094/PDIS-01-20-0203-SC. Epub 2020 Sep 14.
Late blight, caused by the oomycete , is a major constraint on the production of potatoes and tomatoes as well as a constant threat to global food security. An early diagnostic tool is important for the effective management of late blight in the field. Here, in combination with a simplified DNA extraction method, we developed a lateral flow strip-based recombinase polymerase amplification (LF-RPA) assay for the rapid, equipment-free detection of . . This assay targets the Ras-related protein () gene and can be performed over a wide range of temperatures (25 to 45°C). All 12 . isolates yielded positive detection results using the LF-RPA assay, and no cross-reaction occurred with related oomycetes or fungal species. With this assay, the detection limit was 500 fg of genomic DNA in optimized conditions. Furthermore, by combining a simplified polyethylene glycol-NaOH method for extracting DNA from plant samples, the entire LF-RPA assay enabled the detection of within 30 min with no specialized equipment. When applied to field samples, it successfully detected . in naturally diseased potato plants from eight different fields in China. Therefore, the LF-RPA assay is simple, rapid, and cost-effective and has potential for further development as a kit for diagnosing late blight in resource-limited settings or even on-site.
晚疫病由卵菌引起，是马铃薯和番茄生产的主要制约因素，也是全球粮食安全的持续威胁。早期诊断工具对于田间晚疫病的有效管理非常重要。在这里，我们结合简化的 DNA 提取方法，开发了一种基于侧向流动条的重组酶聚合酶扩增（LF-RPA）检测方法，用于快速、无设备检测 。该检测方法针对 Ras 相关蛋白（）基因，可在较宽的温度范围内（25 至 45°C）进行。使用 LF-RPA 检测方法，所有 12 个. 分离株均产生阳性检测结果，与相关卵菌或真菌种无交叉反应。在优化条件下，该检测方法的检测限为 500 fg 基因组 DNA。此外，通过结合一种从植物样本中提取 DNA 的简化聚乙二醇-NaOH 方法，整个 LF-RPA 检测方法可在无专用设备的情况下在 30 分钟内完成对 的检测。当应用于田间样本时，它成功地在来自中国八个不同地区的自然患病马铃薯植株中检测到了 。因此，LF-RPA 检测方法简单、快速且具有成本效益，有潜力进一步开发为资源有限环境甚至现场诊断晚疫病的试剂盒。
