title: Novel approach for the development of axenic microalgal cultures from environmental samples.
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.
authors.affiliation: Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon, 305-806, Korea.
authors.forename: Dae-Hyun
authors.initials: DH
authors.lastname: Cho
authors.affiliation: Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon, 305-806, Korea.
authors.forename: Rishiram
authors.initials: R
authors.lastname: Ramanan
authors.affiliation: Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon, 305-806, Korea.
authors.forename: Byung-Hyuk
authors.initials: BH
authors.lastname: Kim
authors.affiliation: Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon, 305-806, Korea.|Green Chemistry and Environmental Biotechnology, University of Science and Technology, 113 Gwahang-ro, Yuseong-gu, Daejeon, 305-333, Korea.
authors.forename: Jimin
authors.initials: J
authors.lastname: Lee
authors.affiliation: Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon, 305-806, Korea.|Green Chemistry and Environmental Biotechnology, University of Science and Technology, 113 Gwahang-ro, Yuseong-gu, Daejeon, 305-333, Korea.
authors.forename: Sora
authors.initials: S
authors.lastname: Kim
authors.affiliation: Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon, 305-806, Korea.|Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, 305-701, Korea.
authors.forename: Chan
authors.initials: C
authors.lastname: Yoo
authors.affiliation: Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, 305-701, Korea.
authors.forename: Gang-Guk
authors.initials: GG
authors.lastname: Choi
authors.affiliation: Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon, 305-806, Korea.|Green Chemistry and Environmental Biotechnology, University of Science and Technology, 113 Gwahang-ro, Yuseong-gu, Daejeon, 305-333, Korea.
authors.forename: Hee-Mock
authors.initials: HM
authors.lastname: Oh
authors.affiliation: Environmental Biotechnology Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon, 305-806, Korea.|Green Chemistry and Environmental Biotechnology, University of Science and Technology, 113 Gwahang-ro, Yuseong-gu, Daejeon, 305-333, Korea.
authors.forename: Hee-Sik
authors.initials: HS
authors.lastname: Kim
keywords: Axenic culture; Chlorella sp.; FACS; Microalgae; Micropicking; Scenedesmus sp.; Ultrasonication