Skip to main content
SpringerLink
Log in

Isolation of several anti-stress genes from the halotolerant green alga Chlamydomonas by simple functional expression screening with Escherichia coli

  • Published:
World Journal of Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

To isolate anti-stress genes from a halotolerant marine green alga, a simple screening method based on the bacterial expression system was examined. The method consisted of three steps: (i) directional cDNA library construction in a λZAPII vector, (ii) in vivo mass excision of a λZAPII library into phagemid DNA, and (iii) screening for anti-salt-stress and anti-oxidative-stress genes by culturing bacterial cells carrying the in vivo excised phagemid on selection agar plates with a high concentration of NaCl and/or methyl viologen (MV), and by isolating stress-tolerant bacterial colonies. By this method, we screened the cDNA library of halotolerant Chlamydomonas sp. strain W-80, and isolated several stress-related gene homologs, such as glutathione peroxidase, ascorbate peroxidase, bbc1 (breast basic conserved; a low temperature induced gene in Brassica napus), and cyanide-resistant alternative oxidase.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  • Aono, M., Kubo, A., Saji, H., Tanaka, K. & Kondo, N. 1993 Enhanced tolerance to photooxidative stress of transgenic Nicotiana tabacum with high chloroplastic glutathione reductase activity. Plant and Cell Physiology 34, 129-135.

    Google Scholar 

  • Asada, K. 1992 Ascorbate peroxidase-a hydrogen peroxide-scavenging enzyme in plants. Physiologia Plantarum 85, 235-241.

    Google Scholar 

  • Asada, K. 1994 Production and action of active oxygen species in photosynthetic tissues. In Causes of Photooxidative Stress and Amelioration of Defense Systems in Plants, eds Foyer, C. & Mullineaux, P. pp. 77-104. Boca Raton, FL: CRC Press. ISBN 0-84935443-9.

    Google Scholar 

  • Beeor-Tzahar, T., Ben-Hayyim, G., Holland, D., Faltin, Z. & Eshdat, Y. 1996 A stress-associated citrus protein is a distinct plant phospholipid glutathione peroxidase. FEBS letters 366, 151-155.

    Google Scholar 

  • Boeck, A., Forchhammer, K., Heider, J. & Baron, C. 1991 Seleno-protein synthesis: An expansion of the genetic code. Trends in Biochemical Science 16, 463-467.

    Google Scholar 

  • Chomczynski, P. & Sacchi, N. 1987 Single-step method of RNA isolation by acid guanidium thiocyanate-phenol-chloroform extraction. Analytical Biochemistry 162, 156-159.

    Google Scholar 

  • Close, T.J., Kortt, A.A. & Chandler, P.M. 1989 A cDNA-based comparison of dehydration-induced proteins (dehydrins) in barley and corn. Plant Molecular Biology 13, 95-108.

    Google Scholar 

  • Crique, M.C., Jamet, E., Parmentier, Y., Marbach, J., Durr, A. & Fleck, J. 1992 Isolation and characterization of plant cDNA showing homology to animal glutathione peroxidase. Plant Molecular Biology 18, 623-627.

    Google Scholar 

  • Droth, M.P., Duby F., Remacle, C., Dinant, M., Wolwertz, M.R., Duyckaerts, C., Sluse, F. & Matagne, R.F. 1995 Mutations affecting the mitochondrial genes encoding the cytochrome oxidase subunit I and apocytochrome b of Chlamydomonas reinhardtii. Molecular and General Genetics 249, 179-184.

    Google Scholar 

  • Dure, L.I., Crouch, M., Harada, J., Ho, T.H.D., Mundy, J., Quatrano, R., Thomas, T. & Sung, Z.R. 1989 Common amino acid sequence domains among the LEA proteins of higher plants. Plant Molecular Biology 12, 475-486.

    Google Scholar 

  • Eshdat, Y., Holland, D., Faltin, Z. & Ben-Hayyim, G. 1997 Plant glutathione peroxidase. Physiologia Plantarum 100, 234-240.

    Google Scholar 

  • Franzen, L.G., Rochaix, J.D. & Von Heijne, G. 1990 Chloroplast transit peptides from the green alga Chlamydomonas reinhardtii share features with both mitochondrial and higher plant chloroplast presequences. FEBS Letters 260, 165-168.

    Google Scholar 

  • Gupta, A.S., Heinen, J.L., Holaday, A.S., Burke, J.J. & Allen, R.D. 1993 Increased resistance to oxidative stress in transgenic plants that overexpress chloroplastic Cu/Zn superoxide dismutase. Proceedings of National Academy of Science of the USA 90, 1629-1633.

    Google Scholar 

  • Helps, N.R., Adams, S.M., Brammar, W.J. & Varley, J.M. 1995 The Drosophila melanogaster homologue of the human BBC1 is highly expressed during embryogenesis. Gene 162, 245-248.

    Google Scholar 

  • Holland, D., Ben-Hayyim, G., Faltin, Z., Camoin, L., Strosberg, A.D. & Eshdat, Y. 1993 Molecular characterization of salt-stress-associated protein in citrus: protein and cDNA sequence homology to mammalian glutathione peroxidase. Plant Molecular Biology 21, 923-927.

    Google Scholar 

  • Ishikawa, T., Sakai, K., Yoshimura, K., Takeda, T. & Shigeoka, S. 1996 cDNA encoding spinach stromal and thylakoid-bound ascorbate peroxidase, differing in the presence or absence of their 3′-coding regions. FEBS Letters 384, 289-293.

    Google Scholar 

  • Maniatis, T., Fritsch, E.F. & Sambrook, J. 1982 Molecular Cloning, A Laboratory Manual, Cold Spring Harbor, NY: Cold Spring Harbor Laboratory. ISBN No. 0-879689-309-6.

    Google Scholar 

  • Miura, Y., Ohta, S., Mano, M. & Miyamoto, K. 1986 Isolation and characterization of a unicellular marine green alga exhibiting high activity in dark hydrogen production. Agricultural and Biological Chemistry 50, 2837-2844.

    Google Scholar 

  • Miyasaka, H., Ohnishi, Y., Akano, T., Fukatsu, K., Mizoguchi, T., Yagi, K., Maeda, I., Ikuta, Y., Matsumoto, H., Shioji, N. & Miura, Y. 1998 Excretion of glycerol by the marine Chlamydomonas sp. strain W-80 in high CO2 cultures. Journal of Fermentation and Bioengineering 85, 123-125.

    Google Scholar 

  • Moore, A.L., Umbach, A.L. & Siedow, J.M. 1995 Structure-function relationship of the alternative oxidase of plant mitochondria: a model of the active site. Journal of Bioengineering and Biomembranes 27, 367-377.

    Google Scholar 

  • Overbaugh, J.M. & Fall, R. 1985 Characterization of a selenium-independent glutathione peroxidase from Euglena gracilis. Plant Physiology 77, 437-442.

    Google Scholar 

  • Pearson, W.R. & Lipman, D.J. 1988 Improved tools for biological sequence comparison. Proceedings of National Academy of Science of the USA 85, 2444-2448.

    Google Scholar 

  • Purvis, A.C. & Shewfelt, R.L. 1993 Dose the alternative pathway ameliorate chilling injury in sensitive plant tissue? Physiologia Plantarum 88, 712-718.

    Google Scholar 

  • Rabinowitch, H.D., Rosen, G.M. & Fridovich, I. 1987 Contrasting fates of the paraquat monocation radical in Escherichia coli and in Dunaliella salina. Archives of Biochemistry and Biophysics 257, 352-356.

    Google Scholar 

  • Saez-Vasquez, J., Raynal, M., Meza-Basso, L. & Delseny, M. 1993 Two related low-temperature-induced genes from Brassica napus are homologous to the human tumor bbc1 (breast basic conserved) gene. Plant Molecular Biology 23, 1211-1221.

    Google Scholar 

  • Siedow, J.N. & Berthold, D.A. 1986 The alternative oxidase: a cyanide-resistant respiratory pathway in higher plants. Physiologia Plantarum 66, 569-573.

    Google Scholar 

  • Shen, B., Jensen, R.G. & Bohnert, H.J. 1997 Increased resistance to oxidative stress in transgenic plants by targeting mannitol biosynthesis to chloroplast. Plant Physiology 113, 1177-1183.

    Google Scholar 

  • Shikanai, T., Takeda, T., Yamaguchi, H., Sano, S., Tomizawa, K., Yokota, A. & Shigeoka, S. 1998 Inhibition of ascorbate peroxidase under oxidative stress in tobacco having bacterial catalase in chloroplast. FEBS Letters 428, 47-51.

    Google Scholar 

  • Sunde, R.A. 1990 Molecular biology of selenoproteins. Annual Review of Nutrition 10, 451-474.

    Google Scholar 

  • Takeda, T., Ishikawa, T. & Shigeoka, S. 1997 Metabolism of hydrogen peroxide by the scavenging system in Chlamydomonas reinhardtii. Physiologia Plantarum 99, 49-55.

    Google Scholar 

  • Takeda, T., Nakano, Y. & Shigeoka, S. 1993 Effects of selenite, CO2 and illumination on the induction of selenium-dependent glutathione peroxidase in Chlamydomonas reinhardtii. Plant Science 94, 81-88.

    Google Scholar 

  • Takeda, T., Yokota, A. & Shigeoka, S. 1995 Resistance of photosynthesis to hydrogen peroxide in algae. Plant and Cell Physiology 36, 1089-1095.

    Google Scholar 

  • Tarczynski, M.C., Jensen, R.G. & Bohnert, H.J. 1993 Stress protection of transgenic tobacco by production of osmolyte mannitol. Science 259, 508-510.

    Google Scholar 

  • Van Cam, W., Capiau, K., Van Montagu, M., Inze, D. & Slooten, L. 1996 Enhancement of oxidative stress tolerance in transgenic tobacco plants overproducting Fe-superoxide dismutase in chloroplast. Plant Physiology 112, 1703-1714.

    Google Scholar 

  • Wagner, A.M. 1995 A role of active oxygen species as second messengers in the induction of alternative oxidase gene expression in Petunia hybrida cells. FEBS Letters 368, 339-342.

    Google Scholar 

  • Winicov, I. 1993 cDNA encoding putative zinc finger motifs from salt-tolerant alfalfa (Medicago sativa L.) cells. Plant Physiology 102, 681-682.

    Google Scholar 

  • Yokota, A., Shigeoka, S., Onishi, T. & Kitaoka, S. 1988 Selenium as inducer of glutathione peroxidase in low-CO2-grown Chlamydomonas reinhardtii. Plant Physiology 86, 649-651.

    Google Scholar 

  • Yoshimura, K., Ishikawa, T., Nakamura, Y., Tamoi, M., Takeda, T., Tada, T., Nishimura, K. & Shigeoka, S. 1998 Comparative study on recombinant chloroplastic and cytosolic ascorbate peroxidase isozymes of spinach. Archives of Biochemistry and Biophysics 353, 55-63.

    Google Scholar 

  • Zimmermann, J., Voss, H., Schwager, C., Stegemann, J., Erfle, H., Stucky, K., Kristensen, T. & Ansorge, W. 1990 A simplified protocol for fast plasmid DNA sequencing. Nucleic Acids Research 18, 1067.

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. The Kansai Electric Power Co., Technical Research Center, 11-20 Nakoji 3-Chome, Amagasaki, Hyogo 661-0974, Japan

    Hitoshi Miyasaka & Haruo Kanaboshi

  2. Kansai Environmental Engineering Center Co. Ltd, 3-5 Adzuchicho 1-Chome Chuo-ku, Osaka, Osaka 541-0052, Japan

    Kazunori Ikeda

Authors
  1. Hitoshi Miyasaka
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Haruo Kanaboshi
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Kazunori Ikeda
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Miyasaka, H., Kanaboshi, H. & Ikeda, K. Isolation of several anti-stress genes from the halotolerant green alga Chlamydomonas by simple functional expression screening with Escherichia coli. World Journal of Microbiology and Biotechnology 16, 23–29 (2000). https://doi.org/10.1023/A:1008982332139

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1008982332139

Search

Navigation