Library preparation and resequencing
The adipose fins of the 70 individuals selected above that were collected non-invasively and preserved in 99% ethanol were used for DNA extraction. DNA extraction was performed using the DNeasy Blood and Tissue kit (Qiagen, Hilden, Germany) following the manufacturer’s protocol. DNA concentrations were measured using the Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific, Massachusetts, USA). Agarose electrophoresis was performed using KANTO ST (Kanto Chemical, Tokyo, Japan) high gel strength (1% agarose with TAE) to confirm the quality of genomic DNA.
Libraries were prepared from the DNA extracts following the Hackflex protocol (Gaio et al., 2022). For indexes, we used UD Index Adapter (Integrated DNA Technologies, Iowa, USA). Concentrations of the DNA library were measured by a Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific) and the KAPA Library Quantification Kit (Kapa Biosystems) for the StepOnePlus Real-Time PCR System (Thermo Fisher Scientific). Library size was measured using an automated gel electrophoresis system (2200 TapeStation high sensitivity D1000 [Agilent] or 4200 TapeStation high sensitivity D1000 [Agilent]). Fragments in the DNA library were sequenced by using a genome sequencer (HiSeq X; Illumina, California, USA) at 151 bp paired ends.