Total genomic DNA was extracted from the muscle sample using the EasyPure® Genomic DNA Kit according to the manufacturer’s instructions (TransGen Biotech Co, Beijing, China). The integrity of the genomic DNA was assessed by 1% agarose gel electrophoresis and the concentration was assessed using NanoDrop (Thermo Fisher Scientific, Waltham, MA, USA). After DNA extraction and quality detection, a sample was used to sequence the complete mitochondrial genome, which was performed on the Illumina Novaseq 6000 platform (Illumina, San Diego, CA, USA) with PE 2 × 150 bp.
Partial sequences of the cox1 gene were amplified using the primer pairs F – 5′-ACTAATCAYAARGATATTGG-3′ and R – 5′-CCAGTAGGAAYAGCAATAAT-3′ modified from Chen et al. (2011 (link)). Each PCR reaction was performed using a total volume of 50 μl containing 1 μl (approximately 100 ng) of genomic DNA, 25 μl of 2 × EasyTaq PCR SuperMix (TransGen Biotech, Beijing, China), and 1 μl of 10 μmol/l forward and reverse primers. The PCR thermocycler program was as follows: 94 °C for 5 min as initial denaturalization, followed by 35 cycles of 94 °C for 30 s, 50 °C for 1 min, 72 °C for 2 min, and a final extension at 72 °C for 10 min. PCR products were purified and sequenced in both directions on an automatic sequencer (ABI PRISM 3730, Boray Biotechnology Co., Ltd., Xiamen, China).
Partial sequences of the cox1 gene were amplified using the primer pairs F – 5′-ACTAATCAYAARGATATTGG-3′ and R – 5′-CCAGTAGGAAYAGCAATAAT-3′ modified from Chen et al. (2011 (link)). Each PCR reaction was performed using a total volume of 50 μl containing 1 μl (approximately 100 ng) of genomic DNA, 25 μl of 2 × EasyTaq PCR SuperMix (TransGen Biotech, Beijing, China), and 1 μl of 10 μmol/l forward and reverse primers. The PCR thermocycler program was as follows: 94 °C for 5 min as initial denaturalization, followed by 35 cycles of 94 °C for 30 s, 50 °C for 1 min, 72 °C for 2 min, and a final extension at 72 °C for 10 min. PCR products were purified and sequenced in both directions on an automatic sequencer (ABI PRISM 3730, Boray Biotechnology Co., Ltd., Xiamen, China).
