Mitochondrial DNA amplicon sequencing of Trichosurus vulpecula:City_207
The mitochondrial genome was amplified using five overlapping primer pairs. The 5 PCR products were pooled in equivolume amounts and cleaned-up using a 0.8X ratio of SPRI beads diluted in standard PEG buffer (18 % w/v polyethylene glycol 8000 (PEG), 1 M NaCl, 10 mM Tris (pH 8.0), 1 mM EDTA, 0.05 % v/v Tween-20). The concentration of the resulting PCR pool was determined using the Qubit DNA High Sensitivity Kit and Qubit Fluorometer (Invitrogen) and 50 ng of the PCR pool was used to make a DNA library with the NEBNext Ultra II FS DNA Library Prep Kit for Illumina (1/4 reactions; New England Biolabs).