Abstract
Genetic transformation ofStreptomyces caespitosus by plasmid pIj 702 was carried out. Optimal conditions for the protoplast preparation ofStreptomyces caespitosus, its regeneration, and its transformation by pIj 702 were evaluated. Addition of 2% glycine to the culture broth was optimal for protoplast yield. Formation and regeneration of protoplasts were most efficient when the mycelium were harvested at between late log and stationary growth phase. The regeneration frequency of the protoplasts was 15% when the protoplasts were regenerated on R2YE agar media containing 0.5M sucrose. Under the best condition for protoplats regeneration, the optimal transformation frequency was achieved with 40% polyethylene glycol (M.W. 4,000) treatment for 2 minutes.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References Cited
Anderson, M. M., Kibby, J. J., Richards, R. W. and Rothschild, J. M., Biosynthesis of the mitomycin antibiotics from 3-amino-5-hydroxybenzoic acid.J. Chem. Soc., Chem. Commun., 1980, 1277–1278 (1980).
Baltz, R. H., Genetic recombination inStreptomyces fradiae by protoplast fusion and cell regeneration.J. gen. Microbiol., 107, 93–102 (1978).
Epp, J. K., Huber, M. L. B., Turner, J. R. and Schoner, B., Molecular cloning and expression of carbomycin biosynthetic and resistance genes fromStreptomyces thermotolerans: InProceedings of Seventh International Symposium on Biology of Actinomycetes. Japan Scientific Societies Press, Tokyo, 1988, pp. 82–85.
Ghisalba, O., Fuhrer, H., Richter, W. J. and Moss, S., A genetic approach to the rifamycin chromophore inNocardia mediterranei: Isolation and characterization of an early aromatic ansamycin precursor containing the seven-carbon amino starter-unit and three initial acetate/propionate-units of the ansa chain.J. Antibiotics, 34, 58–71 (1981).
Hata, T., Sano, Y., Sugawara, R., Matsumae, A., Kanamori, K., Shina, T. and Moshi, T., Mitomycin, a new antibiotic fromStreptomyces.J. Antibiotics, 9A, 141–149 (1956).
Hopwood, D. A., Genetic studies with bacterial protoplasts.Ann. Rev. Microbiol., 35, 237–272 (1981).
Hopwood, D. A., Bibb, M. J., Chater, K. F., Kieser, T., Ward, C. J. and Schrempf, H., Genetic manipulation ofStreptomyces: A laboratory manual, The John Innes Foundation, Norwich, England, 1985.
Horneman, U., Kehrer, J. P. and Eggert, J. H., Pyruvic acid and D-glucose as precursors in mitomycin biosynthesis byStreptomyces verticillatus.J. Chem. Soc., Chem. Commun., 1974, 1045–1046 (1974).
Katz, E., Thompson, C. J. and Hopwood, D. A. Cloning and Expression of the tyrosinase gene fromStreptomyces antibioticus inStreptomyces lividans.J. Gen. Microbiol., 129, 2703–2714 (1983).
Kiser, T., Factors affecting the isolation of CCC DNA fromStreptomyces lividans andEscherichia coli.Plasmid, 12, 19–36 (1984).
Kim, C. G., Kirschning, A., Bergon, P., Ahn, Y., Wang, J. J., Shibuya, M. and Floss, H. G., 1992. Formation of 3-Amino-5-hydroxybenzoic acid, the precursor of mC7N units in ansamycin antibiotics, by a new variant of the shikimate pathway.J. Am. Chem. Soc., 114, 4941–4943 (1992).
Matsushima, P. and Baltz, R. H., Efficient plasmid transformation ofStreptomyces ambofaciens andStreptomyces fradiae protoplasts.J. Bacteriol., 163, 180–185 (1985).
Nakamura, Y., Ono, E., Kohda, T. and Shinai, H., Highly targeted screening system for carbapenem antibiotics.J. Antibiot, 33, 73–83 (1988).
Okanishi, M., Suzuki, K. and Umezawa, H., Formation and reversion ofStreptomyces protoplasts: Cultural conditions and morphological study.J. Gen. Microbiol., 80, 389–400 (1974).
Pigac, J., Hranueli, D., Smokvina, T. and Alacevic, M., Optimal cultural and physiological conditions for handlingStreptomyces rimosus protoplasts.Appl. Environ. Microbiol., 44, 1178–1186 (1982).
Rinehart Jr., K. L., Potgieter, M., Jin, W.-Z. Pearce C. J., Wright, A., Wright, J. L. C., Walter, J. A. and McInnes, A. G., Biosynthtic studies on antibiotics employing stable isotopes: International conference on Trends in Antibiotic Research, Tokyo, Japan, 1982, pp. 171–184.
Shirahama, T., Furumai, T. and Okanishi, M., A modified regeneration method forStreptomyces protoplasts.Agric. Biol. Chem., 45, 1271–1273 (1981).
Shirling, E. B. and Gottlieb, D., Methods for the characterization ofStreptomyces species.Int. J. Syst. Bacteriol., 16, 313–340 (1966).
Yoo, J. C., Hong, S. W. and Hah, Y. C., Protoplast fusion ofStreptomyces tubercidicus.Kor. J. Microbiol., 24, 364–369 (1986).
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Yoo, J.C., Sim, J.B., Kim, S.J. et al. Genetic transformation ofStreptomyces caespitosus . Arch. Pharm. Res. 16, 300–304 (1993). https://doi.org/10.1007/BF02977520
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF02977520