Journal of Microbiology and Biotechnology

  • 제12권1호
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  • Pages.121-129
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  • 2002
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  • 1017-7825(pISSN)
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  • 1738-8872(eISSN)
한국미생물·생명공학회 (The Korean Society for Microbiology and Biotechnology)
권호 표지

Screening of High-Productivity Cell Lines and Investigation of Their Physiology in Chinese Hamster Ovary (CHO) Cell Cultures for Transforming Growth $Factor-{\beta}1$ Production

  • Chun, Gin-Taek (Division of Life Sciences, Kangwon National University) ;
  • Lee, Joo-Buom (Division of Life Sciences, Kangwon National University) ;
  • Nam, Sang-Uk (Department of Science Educations, Kangwon National University) ;
  • Lee, Se-Won (Division of Life Sciences, Kangwon National University) ;
  • Jeong, Yeon-Ho (Division of Food and Biotechnology, Kangwon National University) ;
  • Choi, Eui-Yul (Department of Genetic Engineering, Hallym University) ;
  • Kim, Ik-Hwan (Graduated School of Biotechnology, Korea University) ;
  • Jeong, Yong-Seob (Division of Biotechnology, Chonbuk National University) ;
  • Kim, Pyeong-Hyeun (Division of Life Sciences, Kangwon National University)
  • 발행 : 2002.02.01
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초록

Using recombinant Chinese hamster ovary (CHO) cells, strategies for developing high producers for the recombinant human Transforming Growth $Factor-{\beta}1$ ($TGF-{\beta}1$) protein are proposed and their physiological characteristics in cell cultures were investigated. $TGF-{\beta}1$ is a pleiotrophic polypeptide involved in various biological activities, including cell growth, differentiation, and deposition of extracellular matrix proteins. The CHO cells included human $TGF-{\beta}1$ cDNA in conjunction with a dihydrofolate reductase (DHFR) gene, which was cotransfected into the cells to amplify the transfected $TGF-{\beta}1$ cDNA. As a first-round screening of the transfected cells, a relatively high $TGF-{\beta}1$-producing cell line was selected, and then, it acquired a resistance to increasing concentrations of methotrexate (MTX) up to $60{\mu}M$,resulting in a significant improvement in its $TGF-{\beta}1$ biosynthetic ability. After applying a monoclonal selection strategy to the MTX-resistant cells, more productive cells were screened, including the APP-3, App-5, and App-8 cell lines. These high producers were compared with two other cell lines (AP-l cell line without amplification of transfected $TGF-{\beta}1$ cDNA and nontransfectant of $TGF-{\beta}1$ cDNA) in terms of cell growth, $TGF-{\beta}1$ productivity, sugar uptake, and byproduct formation, in the presence or absence of MTX in the culture medium. Consequently, both monoclonal selection as well as an investigation of the physiological characteristics were found to be needed for the efficient screening of higher $TGF-{\beta}1$ producers, even after the transfection and amplification of the transfected gene.

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