A Method for the High Yield Preparation of and High Frequency Regeneration of Basidiomycete, Pleurotus ostreatus ("Hiratake"), Protoplasts Using Sulfite Pulp Waste Components

Abstract

An efficient method for the preparation of Pleurotus ostreatus protoplasts (more than 10⁸/ml/2hr) and for colony regeneration from the protoplasts (more than 10%) was developed by adding the components of sulfite pulp wastes to the media and by using an enzyme mixture of Cellulase Onozuka RS, chitinase, Zymolyase 100, 000 and β-glucuronidase. Several different specimens of sulfite pulp waste derivatives (including commercially available ones) stimulated the growth of mycelia suitable as protoplast sources, and markedly increased the regeneration frequency of the protoplasts. The colonies of the regenerated protoplasts could develop fruit bodies effectively within 3 weeks cultivation on media supplemented with the sulfite pulp waste components. The effective components in sulfite pulp wastes were discussed with reference to the studies of K. Inaba et al. and N. Kawamura et al.