Department of Applied Chemistry, Faculty of Science and Technology, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Yokohama, Kanagawa, 223-8522, Japan.
Environmental Management Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), 16-1 Onogawa, Tsukuba, Ibaraki, 305-8569, Japan.
Anal Sci. 2024 Mar;40(3):501-510. doi: 10.1007/s44211-023-00482-7. Epub 2023 Dec 23.
DNA analysis in water samples is attracting attention in various fields. However, conventional methods for DNA analysis require a work-intensive and time-consuming sample pre-treatment. In this study, a simplified pre-treatment method for analyzing DNA in water samples was evaluated. The process consists of filtration, DNA extraction, and amplification, which can be achieved within a short time. In the filtration process, two types of filters, firstly a tissue paper (Kimwipe) and then a glass filter (GF/F), were used in sequence. The first large pore size filter enabled a reduction in filtration time by removing large particulate matter impurities present in river water matrix. Cells spiked into 1 L of river water were recovered at more than 90% within approximately 5 min filtration time. Also, DNA was extracted from the captured cells directly on the surface of the filter in only 5 min. Thus, DNA collection and extraction from a water sample can be completed within about 10 min. Furthermore, PCR amplification was performed directly from DNA-attached filter sections, which greatly reduced the number of required pre-treatment steps. Finally, we succeeded in establishing a simple and fast on-site pre-treatment system by using a hand-driven syringe filtration method. This pre-treatment system is expected to offer the possibility for the future establishment of a rapid and easy DNA analysis method applicable to various types of water samples.
水样中的 DNA 分析在各个领域引起了关注。然而，传统的 DNA 分析方法需要进行繁琐且耗时的样品预处理。本研究评估了一种简化的水样 DNA 分析预处理方法。该过程包括过滤、DNA 提取和扩增，可以在短时间内完成。在过滤过程中，我们先后使用了两种类型的过滤器，首先是纸巾（Kimwipe），然后是玻璃纤维过滤器（GF/F）。第一种大孔径过滤器通过去除河流水体基质中存在的大颗粒杂质，减少了过滤时间。将细胞注入 1 L 的河水中，在大约 5 分钟的过滤时间内，超过 90%的细胞被回收。此外，还可以直接在滤器表面从捕获的细胞中提取 DNA，只需 5 分钟。因此，从水样中收集和提取 DNA 可以在大约 10 分钟内完成。此外，还可以直接从附有 DNA 的滤器部分进行 PCR 扩增，这大大减少了所需的预处理步骤的数量。最后，我们成功地建立了一种简单快速的现场预处理系统，该系统使用手动驱动的注射器过滤方法。这种预处理系统有望为未来建立一种适用于各种类型水样的快速简便的 DNA 分析方法提供可能性。
