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A Multiplex PCR Method for Rapid Identification of Brachionus Rotifers

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Abstract

Cryptic species are increasingly being recognized in many organisms. In Brachionus rotifers, many morphologically similar yet genetically distinct species/biotypes have been described. A number of Brachionus cryptic species have been recognized among hatchery strains. In this study, we present a simple, one-step genetic method to detect the presence of those Brachionus sp. rotifers that have been found in hatcheries. With the proposed technique, each of the B. plicatilis sensu stricto, B. ibericus, Brachionus sp. Nevada, Brachionus sp. Austria, Brachionus sp. Manjavacas, and Brachionus sp. Cayman species and/or biotypes can be identified with polymerase chain reaction (PCR) analysis. Based on 233 cytochrome c oxidase subunit I sequences, we reviewed all the available cryptic Brachionus sp. genetic polymorphisms, and we designed six nested primers. With these primers, a specific amplicon of distinct size is produced for every one of the involved species/biotypes. Two highly sensitive protocols were developed for using the primers. Many of the primers can be combined in the same PCR. The proposed method has been found to be an effective and practical tool to investigate the presence of the above six cryptic species/biotypes in both individual and communal (bulk) rotifer deoxyribonucleic acid extractions from hatcheries. With this technique, hatchery managers could easily determine their rotifer composition at the level of cryptic species and monitor their cultures more efficiently.

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Acknowledgments

This work was supported by an EU research project (ROTIGEN, Q5RS-2002-01302). We thank Dr. N. Karaiskou for her useful comments on multiplexing and all hatcheries and laboratories for providing samples. Finally, we thank Prof. Burgess and two anonymous reviewers for their constructive criticism.

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Authors and Affiliations

  1. Department of Genetics, Development and Molecular Biology, Aristotle University of Thessaloniki, 541 24, Thessaloniki, Greece

    Kalliopi Vasileiadou, Spiros Papakostas, Alexander Triantafyllidis, Ilias Kappas & Theodore J. Abatzopoulos

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  1. Kalliopi Vasileiadou
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  2. Spiros Papakostas
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  3. Alexander Triantafyllidis
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  4. Ilias Kappas
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  5. Theodore J. Abatzopoulos
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Correspondence to Alexander Triantafyllidis.

Additional information

Kalliopi Vasileiadou and Spiros Papakostas contributed equally to this work.

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Vasileiadou, K., Papakostas, S., Triantafyllidis, A. et al. A Multiplex PCR Method for Rapid Identification of Brachionus Rotifers. Mar Biotechnol 11, 53–61 (2009). https://doi.org/10.1007/s10126-008-9119-4

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  • DOI: https://doi.org/10.1007/s10126-008-9119-4

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