Comparing the accuracy and efficiency of third generation sequencing technologies, Oxford Nanopore Technologies, and Pacific Biosciences, for DNA barcode sequencing applications

New genomic technologies, such as third generation sequencing (TGS), have enabled for high-throughput, rapid, and cost-effective data generation of non-model organisms, accelerating taxonomic identification studies and contributing to conservation applications. Here, we present the first comparison of the two leading TGS providers, Pacific Biosciences (PacBio) and Oxford Nanopore Technologies (ONT), for the purpose of DNA barcoding, using morphological identification and Sanger sequencing as the reference. We demonstrate that the highest numbers of successfully sequenced samples were achieved with the ONT R10 & Q20+ chemistry combination. In terms of library preparation time, ONT protocols were the quickest. Compared to Sanger sequencing, we estimate that third-generation platforms become more cost-effective when a study requires the barcoding of more than 61 (Flongle), 183 (MinION), or 356 (PacBio) samples. Although both tested platforms are suitable for DNA barcoding, both had limitations, and applicability to different studies. The pipeline we have developed, which goes from whole specimens to final DNA barcode sequences, can aid planning and budgeting for biodiversity studies, maximising the number of specimens sequenced in one run and speeding up sample processing time.