Introducing ribosomal tandem repeat barcoding for fungi
Journal article, 2019

Sequence comparison and analysis of the various ribosomal genetic markers are the dominant molecular methods for identification and description of fungi. However, new environmental fungal lineages known only from DNA data reveal significant gaps in our sampling of the fungal kingdom in terms of both taxonomy and marker coverage in the reference sequence databases. To facilitate the integration of reference data from all of the ribosomal markers, we present three sets of general primers that allow for amplification of the complete ribosomal operon from the ribosomal tandem repeats. The primers cover all ribosomal markers: ETS, SSU, ITS1, 5.8S, ITS2, LSU and IGS. We coupled these primers successfully with third-generation sequencing (PacBio and Nanopore sequencing) to showcase our approach on authentic fungal herbarium specimens (Basidiomycota), aquatic chytrids (Chytridiomycota) and a poorly understood lineage of early diverging fungi (Nephridiophagidae). In particular, we were able to generate high-quality reference data with Nanopore sequencing in a high-throughput manner, showing that the generation of reference data can be achieved on a regular desktop computer without the involvement of any large-scale sequencing facility. The quality of the Nanopore generated sequences was 99.85%, which is comparable with the 99.78% accuracy described for Sanger sequencing. With this work, we hope to stimulate the generation of a new comprehensive standard of ribosomal reference data with the ultimate aim to close the huge gaps in our reference datasets.

ribosomal operon



third-generation sequencing




Christian Wurzbacher

University of Gothenburg

Technical University of Munich

Gothenburg Global Biodiversity Centre

Ellen Larsson

Gothenburg Global Biodiversity Centre

University of Gothenburg

Johan Bengtsson-Palme

University of Gothenburg

University of Wisconsin Madison

Silke Van den Wyngaert

Leibniz-Institute of Freshwater Ecology and Inland Fisheries

Sten Svantesson

Gothenburg Global Biodiversity Centre

University of Gothenburg

Erik Kristiansson

Chalmers, Mathematical Sciences, Applied Mathematics and Statistics

University of Gothenburg

Maiko Kagami

Leibniz-Institute of Freshwater Ecology and Inland Fisheries

Toho University

Yokohama National University

R. Henrik Nilsson

University of Gothenburg

Gothenburg Global Biodiversity Centre

Molecular Ecology Resources

1755-098X (ISSN) 1755-0998 (eISSN)

Vol. 19 1 118-127

Subject Categories

Biological Systematics

Bioinformatics (Computational Biology)

Bioinformatics and Systems Biology





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