In this research, we aim to select optimal primers facing different aquatic ecosystems. We selected five different study areas, representing five typical aquatic ecosystems (i.e., lakes, marine, wetlands, reservoirs, and rivers). Six sets of parallel samples were collected at the same time and location in each study area. Based on the eDNA method, six common fish universal primers (i.e. 12SRNA, 16SRNA, 18rRNA, MiFish, Cytb, and COI) were selected to detect fish diversity in water samples collected from the same research area.