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Introducing ribosomal tandem repeat barcoding for fungi

Christian Wurzbacher, Ellen Larsson, Johan Bengtsson-Palme, Silke Van den Wyngaert, Sten Svantesson, Erik Kristiansson, Maiko Kagami, R. Henrik Nilsson
doi: https://doi.org/10.1101/310540
Christian Wurzbacher
Department of Biological and Environmental Sciences, University of Gothenburg, Box 461, 40530 Göteborg, Sweden.Chair of Urban Water Systems Engineering, Technical University of Munich, Am Coulombwall 3, Garching 85748, GermanyGothenburg Global Biodiversity Centre, Box 461, 405 30 Göteborg, Sweden
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Ellen Larsson
Department of Biological and Environmental Sciences, University of Gothenburg, Box 461, 40530 Göteborg, Sweden.Gothenburg Global Biodiversity Centre, Box 461, 405 30 Göteborg, Sweden
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Johan Bengtsson-Palme
Wisconsin Institute for Discovery, University of Wisconsin-Madison, 330 North Orchard Street, Madison WI 53715, Wisconsin, USA.Department of Infectious Diseases, Institute of Biomedicine, The Sahlgrenska Academy, University of Gothenburg, Guldhedsgatan 10, 413 46, Göteborg, Sweden
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Silke Van den Wyngaert
Leibniz-Institute of Freshwater Ecology and Inland Fisheries Berlin, Alte Fischerhütte 2, 16775 Stechlin, Germany
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Sten Svantesson
Department of Biological and Environmental Sciences, University of Gothenburg, Box 461, 40530 Göteborg, Sweden.Gothenburg Global Biodiversity Centre, Box 461, 405 30 Göteborg, Sweden
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Erik Kristiansson
Department of Mathematical Sciences, Chalmers University of Technology and University of Gothenburg, 412 96 Göteborg, Sweden
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Maiko Kagami
Leibniz-Institute of Freshwater Ecology and Inland Fisheries Berlin, Alte Fischerhütte 2, 16775 Stechlin, GermanyDepartment of Environmental Science, Faculty of Science, Toho University, 2-2-1 Miyama, Funabashi, Chiba, JapanGraduate School of Environmental and Information Sciences, Yokohama National University, Tokiwadai 797, Hodogayaku, Yokohama, Kanagawa, Japan
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R. Henrik Nilsson
Department of Biological and Environmental Sciences, University of Gothenburg, Box 461, 40530 Göteborg, Sweden.Gothenburg Global Biodiversity Centre, Box 461, 405 30 Göteborg, Sweden
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Abstract

Sequence analysis of the various ribosomal genetic markers is the dominant molecular method for identification and description of fungi. However, there is little agreement on what ribosomal markers should be used, and research groups utilize different markers depending on what fungal groups are targeted. New environmental fungal lineages known only from DNA data reveal significant gaps in the coverage of the fungal kingdom both in terms of taxonomy and marker coverage in the reference sequence databases. In order to integrate references covering all of the ribosomal markers, we present three sets of general primers that allow the 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) from the 5’ end of the ribosomal operon all the way to the 3’ end. We coupled these primers successfully with third generation sequencing (PacBio and Nanopore sequencing) to showcase our approach on authentic fungal herbarium specimens. 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 need for a 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.

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The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted April 28, 2018.
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Introducing ribosomal tandem repeat barcoding for fungi
Christian Wurzbacher, Ellen Larsson, Johan Bengtsson-Palme, Silke Van den Wyngaert, Sten Svantesson, Erik Kristiansson, Maiko Kagami, R. Henrik Nilsson
bioRxiv 310540; doi: https://doi.org/10.1101/310540
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Introducing ribosomal tandem repeat barcoding for fungi
Christian Wurzbacher, Ellen Larsson, Johan Bengtsson-Palme, Silke Van den Wyngaert, Sten Svantesson, Erik Kristiansson, Maiko Kagami, R. Henrik Nilsson
bioRxiv 310540; doi: https://doi.org/10.1101/310540

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