Environmental DNA measurement has been widely applied in organism biomonitoring. Different DNA extraction methods may cause changes in yield and stability, resulting in an inaccurate estimation of eDNA, especially when quantitative measurements are performed. This study focused on the DNA extraction method and compared its yield and stability for stream fish and spiked DNA samples. Samples were collected periodically over a year from river and lake water systems and eDNA was spiked into them. The samples were extracted and compared using three methods: using Buffer-AL for initial lysis with the DNeasy Blood and Tissue Kit (Qiagen); using Buffer-ATL for initial lysis and the microfluidic-channel method (BC method). The method using Buffer-ATL in the DNeasy Blood and Tissue Kit showed better stability and a higher yield than the Buffer-AL method. In addition, the BC method, despite being comparatively simple, performed the extraction stably and with relatively high yields. We showed that differences in DNA extraction methods based on the long-term evaluation of eDNA measurements with various methods may cause alterations in DNA yield and stability.

The environmental DNA (eDNA) method, which is widely applied for biomonitoring, is limited to laboratory analysis and processing. In this study, we developed a filtration/extraction component using a microfluidic channel, Biryu-Chip (BC), and a filtration/extraction method, BC method, to minimize the volume of the sample necessary for DNA extraction and subsequent PCR amplification. We tested the performance of the BC method and compared it with the Sterivex filtration/extraction method using aquarium and river water samples. We observed that using the BC method, the same concentration of the extracted DNA was obtained with 1/20–1/40 of the filtration volume of the Sterivex method, suggesting that the BC method can be widely used for eDNA measurement. In addition, we could perform on-site measurements of eDNA within 30 min using a mobile PCR device. Using the BC method, filtration and extraction could be performed easily and quickly. The PCR results obtained by the BC method were similar to those obtained by the Sterivex method. The BC method required fewer steps and therefore, the risk of DNA contamination could be reduced. When combined with a mobile PCR, the BC method can be applied to easily detect eDNA within 30 min from a few 10 mL of the water sample, even on-site.