Abstract
Factors affecting the PEG-mediated transformation and electrotransformation ofStreptomyces avermitilis protoplasts, an industrial avermectin high-producer, were evaluated. The maximum protoplast transformation efficiency under optimum conditions with PEG was 3 × 106 transformants per µg plasmid pIJ702 DNA. The efficiency of electrotransformation with the same plasmid the intact cells grown in medium with 0.5 mmol/L CaCl2, suspended in buffer with 0.5 mol/L sucrose +1 mmol/L MgCl2, and pulsed at an electric field strength of 10 kV/cm, 800 Θ, 25 µF, was of 2 × 103 transformants per µg DNA. When the cells were electroporated after mild lysozyme-treatment, the efficiency was up to 104 transformants per µg DNA. Electroporation of protoplasts and germlings had a lower efficiency (102 transformants per µg DNA). We report that electroporation under optimum conditions can be used for direct transfer of nonconjugative plasmid pIJ699 between two differentStreptomyces species,S. avermitilis andS. lividans.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
References
Baulard A., Jourdan C., Merchenier A., Locht C.: Rapid mycobacterial plasmid analysis by electroduction betweenMycobacterium spp. andEscherichia coli.Nucl.Acids Res. 20, 4105 (1992).
Bibb B.J., Ward J.M., Hopwood D.A.: Transformation of plasmid DNA intoStreptomyces at high frequency.Nature 274, 398–400 (1978).
Burg R.W., Ostlind D.A., Campbell W.C.: Avermectins, new family of potent anthelmintic agents: producing organism and fermentation.Antimicrob.Agents Chemother. 15, 361–367 (1979).
Harlander S.K.: Transformation ofStreptococcus lactis by electroporation, pp. 229–233 in J.J. Ferretti, R.C. Curtiss III (Eds):Streptococcal Genetics. American Society for Microbiology, Washington (DC) 1987.
Hopwood D.A., Bibb M.J., Chater K.F., Kieser T., Bruton C.J., Kieser H.M., Lydiate D.J., Smith C.P., Ward J.M., Schrempf H.:Genetic Manipulation of Streptomyces,a Laboratory Manual. The John Innes Foundation, Norwich (UK) 1985.
Hwang Y.S., Lee J.Y., Kim E.S., Choi C.Y.: Optimization of transformation procedures in avermectin high-producingStreptomyces avermitilis.Biotechnol.Lett. 23, 457–462 (2001).
Ikeda H., Omura S.: Avermectin biosynthesis.Chem.Rev. 97, 2591–2609 (1997).
Ikeda H., Kotaki H., Tanaka H., Omura S.: Involvement of glucose catabolism in avermectin production byStreptomyces avermitilis.Antimicrob.Agents Chemother. 32, 282–284 (1988).
Ikeda H., Nonomiya T., Omura S.: Organization of the biosynthetic gene cluster for the polyketide anthelmintic macrolide avermectin inStreptomyces avermitilis.Proc.Nat.Acad.Sci.USA 96, 9509–9514 (1999).
Lee J.Y., Hwang Y.S., Kim E.S., Choi C.Y.: Effect of a global regulatory gene,afsR2, fromStreptomyces lividans on avermectin production inStreptomyces avermitilis.J.Biosci.Bioeng. 89, 606–608 (2000).
MacNeil D.J.: Introduction of plasmid DNA intoStreptomyces lividans by electroporation.FEMS Microbiol.Lett. 42, 239–244 (1987).
MacNeil D.J., Klapko L.M.: Transformation ofStreptomyces avermitilis by plasmid DNA.J.Ind.Microbiol. 2, 209–218 (1987).
Matsushima P., Baltz R.H.: Efficient plasmid transformation.J.Bacteriol. 163, 180–185 (1985).
Mazy-Servais C., Baczkowski D., Dusart J.: Electroporation of intact cells ofStreptomyces parvulus andStreptomyces vinaceus.FEMS Microbiol.Lett. 151, 135–138 (1997).
O’Sullivan T.F., Fitzgerald G.F.: Electrotransformation of industrial strains ofStreptococcus thermophilus.J.Appl.Microbiol. 86, 275–283 (1999).
Pfau J., Youderian P.: Transferring plasmid DNA between different bacterial species with electroporation.Nucl.Acids Res. 18, 6165 (1990).
Pigac J., Schrempf H.: Optimal cultural and physiological conditions for handlingStreptomyces rimosus protoplasts.Appl.Environ.Microbiol. 44, 1178–1186 (1982).
Pigac J., Schrempf H.: A simple and rapid method of transformation ofStreptomyces runosus R6 and other streptomycetes by electroporation.Appl.Environ.Microbiol. 61, 352–356 (1995).
Serror P., Sasaki T., Ehrlich S.D., Maguin E.: Electrotransformation ofLactobacillus delbrueckii subsp.bulgaricus andL. delbrueckii subsp.lactis with various plasmids.Appl.Environ.Microbiol. 68, 46–52 (2002).
Shivarova N., Forster W., Jacob H.E., Grigorava R.: Microbiological implications of electrical field effects. VII. Stimulation of plasmid transformation ofBacillus cereus protoplasts by electric field pulses.Z.Allg.Mikrobiol. 23, 595–599 (1983).
Süleymanoğlu E.: Electrorelease ofEscherichia coli nucleoids.Folia Microbiol. 47, 365–370 (2002).
Summers D.K., Withers H.L.: Electrotransfer: direct transfer of bacterial plasmid DNA by electroporation.Nucl.Acids Res. 18, 2192 (1990).
Tyurin M., Starodubtseva L., Livshits V.: Electrotransformation of germinating spores ofStreptomyces spp.Biotech.Tech. 9, 737–740 (1995).
Vujaklija D., Davies J.: Direct transfer of plasmid DNA betweenStreptomyces spp. andE. coli by electroduction.J.Antibiot. 48, 635–637 (1995).
Weaver J.C.: Electroporation protocol for microorganisms, pp. 1–26 in J.A. Nicroloff (Ed.):Methods in Molecular Biology, Vol. 47, Humana Press, Totowa (USA) 1990.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Gong, W., Jiang, W., Yang, Y. et al. Improvement of transformation and electroduction in avermectin high-producer,Streptomyces avermitilis . Folia Microbiol 49, 399–405 (2004). https://doi.org/10.1007/BF02931600
Received:
Revised:
Published:
Issue Date:
DOI: https://doi.org/10.1007/BF02931600