CULTIVATION‐INDEPENDENT SPECIES IDENTIFICATION OF DINOBRYON SPECIES (CHRYSOPHYCEAE) BY MEANS OF MULTIPLEX SINGLE‐CELL PCR1 |
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Authors: | Steffen Jost Ralph Medinger Jens Boenigk |
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Affiliation: | 1. Institute for Limnology, Austrian Academy of Sciences, Mondseestr. 9, 5310 Mondsee, Austria;2. Author for correspondence: e‐mail .;3. Institute for Limnology, Austrian Academy of Sciences, Mondseestr. 9, 5310 Mondsee, Austria Institute for Ecology, University of Innsbruck, Leopold‐Franzens‐University Innsbruck, Technikerstr. 25, 6020 Innsbruck, Austria |
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Abstract: | With the discovery of a high molecular diversity of protists, a discrepancy between morphological and molecular species richness estimates became apparent. Solving the current concerns requires a comparative analysis of different sequences combined with morphological analyses of single cells originating from preserved field samples. We refined a single‐cell PCR (SC‐PCR) protocol for analyzing cells from field samples preserved with Lugol’s iodine solution. We linked microscopic screening with multiplex PCR targeting the SSU rDNA, internal transcribed spacer 1 (ITS1), 5.8S rDNA, internal transcribed spacer 2 (ITS2), and the mitochondrial cytochrome oxidase 1 (CO1) in a single PCR reaction. Using this method, we investigated the intraspecific molecular variation in Dinobryon populations originating from two lakes in the Salzkammergut area of Austria. All investigated genetic markers showed two separated clusters within the investigated populations of Dinobryon divergens O. E. Imhof, indicating a reproductive isolation of the two coexisting populations. Based on these findings, we describe a lineage, which is morphologically similar to D. divergens but, based on the molecular data, is reproductively isolated. |
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Keywords: | chromalveolates chromist Chrysophyceae golden algae multiplex single‐cell PCR taxonomy |
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