Factors affecting the protoplast formation and regeneration of Bacillus pumilus and Cellulomonas fimi for intergeneric protoplast fusion

Several factors predicted to affect the protoplast formation and regeneration were investigated. The optimum lysozyme, casamino acid and PVP concentration were 0.5 (mg/$m\ell$), 0.1 (%) and 1.5(%). In B. pumilus, Penicillin-G treatment concentration was 0.3 (U/$m\ell$) and optimum treatment period was transit log. phase. And in the case of Celm. fimi, 0.3 (U/$m\ell$) and initial log. phase. Osmotic stabilizer and di-cation for OSM medium of B.pumilus and Gelm .fimi were 25mM CaCl2, 0.5M sodium sucinate and 50mM MgCl$_2$, 100mM CaCl$_2$, 0.4M sodium succinate. The regeneration frequency of B.pumilus and Celm. fimi were 14.6(%) and 6.9(%).