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Appl Environ Microbiol. 1982 November; 44(5): 1178-1186
Copyright © 1982, American Society for Microbiology. All Rights Reserved.

Optimal Cultural and Physiological Conditions for Handling Streptomyces rimosus Protoplasts

Jasenka Pigac, Daslav Hranueli, Tamara Smokvina and Marija Alaevi

PLIVA Pharmaceutical, Chemical, Food, and Cosmetic Industry, and Faculty of Food and Biotechnology, University of Zagreb, Zagreb, Yugoslavia

ABSTRACT

A general procedure for manipulating protoplasts of three Streptomyces rimosus strains was developed. More than 50% regeneration efficiency was obtained by optimizing the osmotic stabilizer concentrations and modifying the plating procedure. Preparation and regeneration of protoplasts were studied by both phase-contrast and electron microscopy. After cell wall degradation with lysozyme, protoplasts about 1,000 to 1,500 nm in diameter appeared. The reversion process exhibited normal and aberrant regeneration of protoplasts to hyphae and to spherical cells, respectively. Spherical cells contained no , -LL-diaminopimelic acid and were colorless or red after Gram staining. They showed consistent stability during at least five subsequent subcultivations. However, the omission of glycine from the precultivation medium reduced the unusual process of regeneration almost completely. After normal protoplast regeneration, the production of oxytetracycline by single isolates was not affected.

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