Jessica Rieder, Eliane Jemmi, Irene Adrian-Kalchhauser 

Ecologists, conservation biologists, and environmental managers are increasingly dependent on environmental DNA (eDNA) data for research and decision making. The inherent complexity of eDNA samples, coupled with different choices involved during the DNA extraction process, can introduce biases into the final eDNA dataset. Therefore, optimization and troubleshooting of DNA extraction protocols are pivotal for the successful execution of eDNA projects. Knowledge of the basic steps and principles of DNA extraction is essential for eDNA analysis. However, traditional education in ecology, conservation, and environmental management typically does not include in-depth training in molecular methods. While DNA extraction box kits are typically user-friendly, they may fail to deliver the desired results with eDNA samples, necessitating protocol adaptations or educated selection of alternative approaches.
The primary objective of this paper is to enable scientists with an ecological background who employ DNA extraction protocols to understand the four key steps of DNA extraction, and to use this expertise to their advantage. Furthermore, we describe the purpose of commonly used reagents and chemicals, point out alternatives for each key step, explain the impact of certain choices on DNA integrity and purity, and advocate for an adaptable “mix and match” protocol when applicable.
We anticipate that the paper will enable field ecologists to develop a deeper understanding of the mechanisms and chemistry underlying DNA extraction, thus allowing them to make informed decisions regarding the best DNA extraction method for their research. Our intention is not to provide comprehensive, step-by-step protocols, but to offer guiding principles while highlighting alternative solutions. Finally, we hope that this paper will act as a useful resource to support knowledge transfer and teaching.