DNA barcoding of Pleuronectiformes: in silico analysis and development of markers

DNA barcoding is a method used for the identification and discovery of animal species. It usually involved a 648 base pair fragment of the mitochondrial cytochrome c oxidase subunit I, known as COI. This work is focused on the study of the genetic identification in the families belonging to the order Pleuronectiformes, commonly known as flatfish, and the accuracy of the genetic marker most used for the study of their DNA barcodes. The results indicate possible existence of taxonomical mistakes because several families do not show a gap between maximum intraspecific distance - which is the maximum distance within a specie - and the minimum interspecific distance - which is the minimum distance between a species and its nearest neighbor (NN), meaning that the marker in use cannot reliably distinguish among those species. This study uses a bioinformatic approach to design new Pleuronectiformes barcodes and compares their coverage and resolving power with that of existing barcodes. The new primers, proposed by the program EcoPrimer, are based on two indices that estimate the resolution capacity of the barcodes and the taxonomic coverage of them, for the amplification. The performances of both barcoding regions already in use (COI and 16 rDNA genes), and the new primer pairs designed, were performed through a ‘in silico PCR’. The results show that the new primer pairs, located in a different regions of 16S rDNA gene compared to the universal barcode region used in fishes, present best resolution capacity and taxonomic coverage than the others already in use. This is an essential complement for future barcoding studies.