metabarcoding using CO1 primers
PCR amplification targeted multiple mitochondrial gene markers: CO1, to detect a broad range of manager-relevant fish species in the region. Fluidigm CS1 (forward) and CS2 (reverse) common oligonucleotide tags were added to the 5 ends of primers to support two-step library preparation. PCR was conducted in triplicate for each sample, with pooled amplicons post-PCR. For 25-L reactions (used for MarVer1 and MarVer3): 12.5L Platinum Taq, 8.5L HO, 1L of each 10mM primer, and 2L template DNA. For 12.5-L reactions (used for MiFish): 6.25L Platinum Taq, 4.25L HO, 0.5L of each primer, and 1L template DNA. PCRs followed a touchdown protocol: 95C for 15:00; 13 cycles of 94C for 00:30, 60C for 00:30, 72C for 01:30; 25 cycles of 94C for 00:30, 60C for 00:30, 72C for 00:45; final extension at 72C for 10:00; hold at 4C.