Using next-generation sequencing to improve DNA barcoding: lessons from a small-scale study of wild bee species (Hymenoptera, Halictidae)

• DOI: 10.1007/s13592-018-0594-y
• Publication Date: 2018-08-31T11:49:54Z
• Authors: Gontran Sonet; Alain Pauly; Zoltán T. Nagy; Massimiliano Virg...; Kurt Jordaens; Jeroen Van Houdt; Sebastian Worms; Marc De Meyer; Thierry Backeljau
• Keywords: Sanger sequencing; Halictidae; DNA Barcoding; Amplicon; Ion semiconductor sequencing

The parallel sequencing of targeted amplicons is a scalable application of next-generation sequencing (NGS) that can advantageously replace Sanger sequencing in certain DNA barcoding studies. It can be used to sequence different PCR products simultaneously, including co-amplified products. Here, we explore this approach by simultaneously sequencing five markers (including the DNA barcode and a diagnostic marker of Wolbachia) in 12 species of Halictidae that were previously DNA barcoded using Sanger sequencing. Consensus sequences were obtained from fresh bees with success rates of 74–100% depending on the DNA fragment. They improved the phylogeny of the group, detected Wolbachia infections (in 8/21 specimens) and characterised haplotype variants. Sequencing cost per marker and per specimen (11.43 €) was estimated to decrease (< 5.00 €) in studies aiming for a higher throughput. We provide guidelines for selecting NGS or Sanger sequencing depending on the goals of future studies.