A comparison of DNA barcoding markers in West African frogs

Abstract DNA barcoding has been proposed as a means of quick species identification using a short standardised segment of DNA. The established barcode gene for animals—the mitochondrial gene cytochrome oxidase one (CO1)—has been plagued by primer failure and low species identification success in amphibians. We investigate the accuracy of CO1 barcoding with a new dataset of West African frogs using the universal CO1 primers and new amphibian-specific CO1 primers in comparison to a proposed alternative DNA barcode for amphibians—the mitochondrial ribosomal 16s gene (16s). Research was performed using 134 specimens, comprising 21 species collected in Ghana, a global biodiversity hotspot with a deficiency of amphibian barcoding resources. These species represent 55% of amphibian species (58% of amphibian families) that are known in the area from surveys from 1988 to 2009. We found nearly a 50% increase in PCR amplification success using the amphibian-specific CO1 primers compared to the universal CO1 primers. However, the overall amplification and sequencing success of the amphibian-specific CO1 primers was low (78%) compared to the 16s gene (100%). Neither marker has a clear advantage in terms of barcoding gap; comparisons of intraspecific and interspecific variation for these markers were similar for the species we examined. Considering the qualities a barcoding gene should possess, 16s outperformed CO1 in terms of ease of obtaining sequences, and given that 16s sequences are better represented for African frogs on GenBank, this marker had higher success in BLAST searches. With amphibian species in fast decline, more consideration should be given to the appropriateness of collecting CO1 barcodes for amphibians, especially as an extensive genetic database for 16s already exists that can accurately identify amphibians.