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Validation of Housekeeping Genes for Gene Expression Studies in Symbiodinium Exposed to Thermal and Light Stress

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Abstract

Unicellular photosynthetic algae (dinoflagellate) from the genus Symbiodinium live in mutualistic symbiosis with reef-building corals. Cultured Symbiodinium sp. (clade C) were exposed to a range of environmental stresses that included elevated temperatures (29°C and 32°C) under high (100 μmol quanta m−2 s−1 Photosynthetic Active Radiation) and low (10 μmol quanta m−2 s−1) irradiances. Using real-time RT-PCR the stability of expression for the nine selected putative housekeeping genes (HKGs) was tested. The most stable expression pattern was identified for cyclophilin and S-adenosyl methionine synthetase (SAM) followed by S4 ribosomal protein (Rp-S4), Calmodulin (Cal), and Cytochrome oxidase subunit 1 (Cox), respectively. Thermal stress alone resulted in the highest expression stability for Rp-S4 and SAM, with a minimum of two reference genes required for data normalization. For Symbiodinium exposed to both, light and thermal stresses, at least five reference genes were recommended by geNorm analysis. In parallel, the expression of Hsp90 for Symbiodinium in culture and in symbiosis within coral host (Acropora millepora) was evaluated using the most stable HKGs. Our results revealed a drop in Hsp90 expression after an 18 h-period and a 24 h-period of exposure to elevated temperatures indicating the similar Hsp90 expression profile in symbiotic and non-symbiotic environments. This study provides the first list of the HKGs and will provide a useful reference in future gene expression studies in symbiotic dinoflagellates.

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Acknowledgments

We would like to thank to Dr. Bill Leggat for his work on the Symbiodinium (C3) EST project. Also, thanks to Dr. Olga Pantos, Dr. Shona Osborne, Dr. Deanne Mitchell and Alicia Crawley for their comments on the manuscript draft and Ruth Reef for an egg-sperm sample from A. millepora. This project was supported by ARC Centre for Excellence in Reef Studies, a Marie Curie International Outgoing Fellowship to M.P. and the Coral Reef Targeted Research Project (www.gefcoral.org).

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Authors and Affiliations

  1. Global Change Institute, University of Queensland, St. Lucia, Brisbane, 4072, QLD, Australia

    Nedeljka N. Rosic, Mathieu Pernice & Ove Hoegh-Guldberg

  2. ARC Centre for Excellence in Reef Studies, University of Queensland, St. Lucia, Brisbane, 4072, QLD, Australia

    Ove Hoegh-Guldberg

  3. UMR 7208 “Biologie des Organismes et Ecosytèmes Aquatiques” MNHN-CNRS-IRD–UPMC, Case postale 53, 75231, Paris cedex 05, France

    Mathieu Pernice

  4. Department of Biology, University of Louisiana at Lafayette, Lafayette, LA, 70504, USA

    Mauricio Rodriguez-Lanetty

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  1. Nedeljka N. Rosic
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  3. Mauricio Rodriguez-Lanetty
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  4. Ove Hoegh-Guldberg
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Correspondence to Nedeljka N. Rosic.

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Rosic, N.N., Pernice, M., Rodriguez-Lanetty, M. et al. Validation of Housekeeping Genes for Gene Expression Studies in Symbiodinium Exposed to Thermal and Light Stress. Mar Biotechnol 13, 355–365 (2011). https://doi.org/10.1007/s10126-010-9308-9

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