Nagai Satoshi, Itakura Shigeru
National Research Institute of Fisheries and Environment of Inland Sea, Maruishi 2-17-5, Hatsukaichi, Hiroshima 739-0452, Japan.
Mar Genomics. 2012 Sep;7:43-9. doi: 10.1016/j.margen.2012.03.001. Epub 2012 Apr 5.
In this study, we succeeded in developing a loop-mediated isothermal amplification (LAMP) method that enables sensitive and specific detection of the toxic marine dinoflagellates Alexandrium tamarense and Alexandrium catenella from single cells of both laboratory cultures and naturally blooming cells within 25 min, by monitoring the turbidimeter from the start of the LAMP reaction. The fluorescence intensity was strong enough to allow discrimination between positive and negative results by naked eye under a UV lamp, even in amplified samples from a single cell, by using the LAMP method. Unambiguous detection by naked eye was possible even in half the volume of LAMP cocktail recommended by the manufacturer, suggesting the potential to significantly reduce the cost of Alexandrium monitoring. Therefore, we can conclude that this method is one of the most convenient, sensitive, and cost-effective molecular tools for Alexandrium monitoring.
在本研究中，我们成功开发了一种环介导等温扩增（LAMP）方法，通过在LAMP反应开始时监测浊度仪，能够在25分钟内从实验室培养的单细胞以及自然爆发的细胞中灵敏且特异检测有毒海洋甲藻塔玛亚历山大藻（Alexandrium tamarense）和链状亚历山大藻（Alexandrium catenella）。使用该LAMP方法时，荧光强度足够强，即使在单细胞的扩增样品中，也能在紫外灯下通过肉眼区分阳性和阴性结果。即使使用制造商推荐的LAMP反应液体积的一半，也能够通过肉眼进行明确检测，这表明有可能显著降低亚历山大藻监测的成本。因此，我们可以得出结论，该方法是用于亚历山大藻监测的最便捷、灵敏且经济高效的分子工具之一。
