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It has not been much known about that how much the properties of parental strains appear in the monokaryotic progeny of their basidiospores and the properties of monokaryotic progeny is transferred and expressed after mating with other monokaryotic progeny. To get a basic idea in the evaluation of shiitake strains that are subjectto be used for a breeding program, biochemical properties of monokaryotic strains generated from basidiospores of parental shiitake strains (Sanzo101 and Sanzo108) and hybrid strains generated from cross of these monokaryotic strains were examined. When amylase, avicelase, β-glucosidase, CM-cellulase, pectinase, proteinase, and xylanase were tested against monokaryotic strains generated from forty basidiospores of the two parental strains, No identical patterns of the degree of enzyme activity were observed between monokaryotic strains and parental strains of the two shiitake cultivars. The degree of extracellular enzyme activity also varied among monokaryotic strains of the two shiitake cultivars. Our results showed that dikaryotic parental strains of shiitake mushroom produce basidiospores having very diverse biochemical properties. Biochemical test of monokaryotic strains was useful for the comparative evaluation of parental strains. Biochemical test with hybrid strains formed with the combinational cross of monokaryotic strains that have different degree of extracellular activity showed that the property of monokaryotic strains is not expressed in the hybrid strains according to Mandelian principles. Tests of the ability of using diverse polymeric carbon sources, tolerance to copper and agrochemicals, and antagonistic property to Tricoderma atroviride also showed similar results