Protoplast Formation and Regeneration ofStreptococcus lactisCells

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Abstract
Conditions for protoplast formation and cell wall regeneration of Streptococcus lactis were described. Since S. lactis was relatively resistant to the action of lysozyme, cell wall digestion was insufficient when the cells were treated with lysozyme alone. Protoplasts were obtained by treatment with lysozyme and a bacterial α-amylase preparation in Tris-HCl buffer containing 3 mM MgCl2 and 20% sucrose. The number of remaining osmotically stable cells decreased to less than 9 × 10−7. Regeneration was accomplished with a hypertonic medium that contained CaCl2, MgCl2, gelatin and sucrose. The frequency of regeneration of protoplasts to normal cells was 3 ~ 10% after 5 ~ 7 days incubation in the hypertonic medium.