環境DNA 分析によるグッピーPoecilia reticulataのオプシン遺伝子LWS-1 多型の検出

抄録

Environmental DNA (eDNA)-based analysis has been developed as an effective method for estimating fish fauna and detecting endangered and invasive species in water. Although the major marker genes in fishes and vertebrates are encoded in mitochondrial genomes, other genes on nuclear genome would also be feasible for eDNA studies. To address this possibility, we focused on the opsin LWS-1 (long-wave sensitive opsin-1) gene of guppy Poecilia reticulata. We designed polymerase chain reaction (PCR) primers to amplify approximately 158 base pairs of guppy LWS-1 to distinguish their functional polymorphic alleles LWS-1_Ala (A-type) and LWS-1_Ser (S-type). Using these primers and eDNA samples of Okinawa Island, Japan, we confirmed that the nuclear-encoded opsin gene of guppy was successfully sequenced using the Illumina MiSeq DNA sequencer. The analysis of obtained sequences demonstrated that red-shifted S-type allele was dominated in Senbaru Pond, where the turbidity was stably lower. On the other hand, both S- and A-types were detected in Makiminato River, where the turbidity was relatively high. Such association between functional allele variations of LWS-1 and transparency of living water has also been reported in guppy of Trinidad and Tobago Islands. We suggest that the eDNA-based analysis can be applied to nuclear-encoded genes of fishes unless the biomass of target species is quite low. Our findings provide important advance in understanding molecular evolution and adaptation process in the wild populations of target materials on the basis of meta-genomic approach.