To improve alkaline protease producing strain by protoplast fusion, a strain of Bacillus sp. was treated with 100§¶/§¢ NTG (N-methyl-N¢¥-vitro-N-nitrosoguanidine) for 45 minutes and mutants of Bacillus subtilts Arg^-, Try^-, His^- and Ade^- were isolated. The frequency of protoplast formation was about 99%, when teas of exponential phase were treated with 200§¶/§¢ lysozyme at 42¡É for 10¡30 minutes. In a regeneration medium containing 0.3M sodium succinate, 2.0% polyvinylpyrrolidone, 0.5% casamino acid, 10mM MgCl©ü and 20mM CaCl©ü, regeneration frequency cf the isolated Bacillus subtilis strains was 25.2%. The fusion frequency between mutant, was from 2.1¡¿10^(-5) to 8.1¡¿10^(-5) under optimum condition.
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