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Novel approach for the development of axenic microalgal cultures from environmental samples

Authors

  • Dae-Hyun Cho,

    1. Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
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  • Rishiram Ramanan,

    1. Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
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  • Byung-Hyuk Kim,

    1. Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
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  • Jimin Lee,

    1. Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
    2. Green Chemistry and Environmental Biotechnology, University of Science and Technology, Daejeon, Korea
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  • Sora Kim,

    1. Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
    2. Green Chemistry and Environmental Biotechnology, University of Science and Technology, Daejeon, Korea
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  • Chan Yoo,

    1. Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
    2. Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, Korea
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  • Gang-Guk Choi,

    1. Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, Korea
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  • Hee-Mock Oh,

    1. Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
    2. Green Chemistry and Environmental Biotechnology, University of Science and Technology, Daejeon, Korea
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  • Hee-Sik Kim

    Corresponding author
    1. Green Chemistry and Environmental Biotechnology, University of Science and Technology, Daejeon, Korea
    • Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
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Author for correspondence: e-mail hkim@kribb.re.kr.

Abstract

We demonstrated a comprehensive approach for development of axenic cultures of microalgae from environmental samples. A combination of ultrasonication, fluorescence-activated cell sorting (FACS), and micropicking was used to isolate axenic cultures of Chlorella vulgaris Beyerinck (Beijerinck) and Chlorella sorokiniana Shihira & R.W. Krauss from swine wastewater, and Scenedesmus sp. YC001 from an open pond. Ultrasonication dispersed microorganisms attached to microalgae and reduced the bacterial population by 70%, and when followed by cell sorting yielded 99.5% pure microalgal strains. The strains were rendered axenic by the novel method of micropicking and were tested for purity in both solid and liquid media under different trophic states. Denaturing gradient gel electrophoresis (DGGE) of 16S rRNA gene confirmed the absence of unculturable bacteria, whereas fluorescence microscopy and scanning electron microscopy (SEM) further confirmed the axenicity. This is the most comprehensive approach developed to date for obtaining axenic microalgal strains without the use of antibiotics and repetitive subculturing.

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