A uniparental mitochondrial (mt) transmission pattern has been previously observed in laboratory matings of the cultivated mushroom Agaricus bisporus on petri dishes. In this study, four sets of specific matings were further examined by taking mycelial plugs from the confluent zone of mated homokaryons and inoculating these plugs into rye grain for laboratory fruiting and for fruiting under industrial conditions. Examination of the mt genotype of each individual fruit body for mt-specific restriction fragment length polymorphisms further confirmed that the mt genome was inherited uniparentally. The vegetative radial growth and the fruiting activity of two pairs of intraspecific heterokaryons, each pair carrying the same combination of nuclear genomes but different mt genotypes, were compared. Our results suggested that the mt genotype did not appreciably affect radial growth or fruiting activity. The failure to recover both heterokaryons, each carrying either parental mt genotype in any given cross, therefore clearly indicated that in matings of A. bisporus, the mt genome from one of the parental homokaryons is either selectively excluded in the newly formed heterokaryon or selectively eliminated in the immediate heterokaryotic mitotic progeny of the newly formed heterokaryon.