Martin KAPUN, Bioinformaticien en Génomique au Muséum d'Histoire Naturelle de Vienne

Recent advances in sequencing technologies facilitate simultaneous sequencing of millions of DNA fragments in a highly parallelized and cost-efficient manner. In particular, recently developed third generation sequencing methods allow to sequence long DNA fragments, which makes it possible to simultaneously obtain sequence information from whole PCR amplicons for hundreds of samples that can be used for quantitative DNA barcoding. However, the application of these approaches is still often challenged by high sequencing error rates and complexity of the data, which requires advanced bioinformatics know-how to address these problems. In the project that we conceived as part of the TETTRIs consortium, we aim to alleviate these challenges by developing a simple, yet powerful combination of wet lab and bioinformatic tools to analyse closely related taxa based on multiple PCR markers. Our pipeline allows to reconstruct phased consensus sequences from multiplexed multi-locus PCR amplicons sequenced with Oxford Nanopore technology and provides phylogenetic analysis methods and species identification by comparison to the BOLD database. In this seminar, I will describe the wet lab procedures to optimize multiplexed PCR amplification of multiple nuclear and mitochondrial markers and will demonstrate the application of our approach to the phylogenetic analysis of Austrian Syrphid hoverflies.