Dna sequencing

DNA sequencing

DNA was extracted using the Epmotion 5075 robot (Eppendorf), following the manufacturers’ recommendations. The barcode fragment of the cytochrome oxidase subunit I ( cox1) was amplified using universal primers LCO1490/ HCO2198 ( Folmer et al., 1994), and PCRs wErE pErforMEd In 25 μL, conTaInInG 3 nG of DNA , 1X rEacTIon buffEr, 2.5 mM MgCl2, 0.26 mM dNTP, 0.3 mM each primer, 5% DMSO and 1.5 units of Qbiogene Q-Bio Taq, and the amplification steps consisted of an initial denaturation step at 94 °C for 4 min, followed by 35 cycles of denaturation at 94 °C for 30 s, annealing at 50 °C for 30 s, followed by extension at 72 °C for 1 min. The final extension was at 72 °C for 5 min. PCR products were purified and sequenced by the Eurofins sequencing facility.