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KMID : 0364819860240010024
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Korean Journal of Microbiology
1986 Volume.24 No. 1 p.24 ~ p.31
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The Spheroplast Formation and Regeneration of Pseudomonas spp.
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Abstract
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1
The optimal conditions for the formation and the regeneration of Pseudumunas spheroplast were measured. Pseudomonas spp. cells were transformed to spheroplasts from 99.0% to 99.9% by treatment of 100 ug/ml lysozyme and 10mM EDTA at room temperature. The optimal pH for the spheroplast formation was pH 8.0. Magnecium chloride, calcium chloride and streptomycin were effective on the stabilization of Pseudomonas spheroplast, while Mg + and ZVa * ions were effective on the formation of Pseudomonas spheroplast. Rich Regeneration Medium was used for the regeneration of Pseudomonas spheroplast. To improve regeneration frequency, Bovin Serum Albumine and cationic ions were added to the spheroplast dilution beffer and regeneration environment respectively. Treatment of 20mM calcium chloride in the Rich Regeneration Medium could improve the yield of regenerants as much as 28-fold. Treatment of 1% Bovin Serum Albumine in the spheroplast formation and dilution buffer increased the yield of regenerants to 10-fold. Also, the regeneration frequency was improved to 14-fold when Rich Regeneration Meidtun containing 0.5% Gelatin was used for regeneration as well as 1% Bovin Serum Albumine.
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