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The mating types of 124 and 88 single spore isolates, isolated at random from Volvariella volvacea strains V23 ( A mating type locus alleles A1 + A2 ) and PY ( A mating type locus alleles A3 + A4 ) respectively, were determined by molecular screening using four primer pairs designed according to the sequences of the A mating type genes ( HD1 and HD2 ) of the two parental strains. Of the single spore isolates from strain V23, 101were confirmed as homokaryons, 35 of which were mating type A1 and 66 mating type A2 . The remaining 23 were heterokaryons carrying both A1 and A2 mating type loci. Of the single spore isolates from strain PY, 72 were confirmed as homokaryons, of which 41 were mating type A3 and 31 mating type A4 . Sixteen were heterokaryons carrying both A3 and A4 mating type loci. Cross-breeding between 72 compatible pairs generated 58 hybrids, three of which were designated high quality hybrids based on agronomic traits determined by cultivation experiments. Our research has established that the molecular marker-assisted cross-breeding technique represents a practical and highly efficient system for generating improved V. volvacea cultivars. [ABSTRACT FROM AUTHOR]