The primary objective of the present study is the characterization of the hybrids of dikaryon-monokaryon(di-mono) and monokaryon-monokaryon(mono-mono) crosses in mushroom breeding. We employed this technique for developing develop superior species from Pleurotus spp. varieties with 56 Di-mono intraspecific hybrids of 14 combinations and 85 mono-mono intraspecific hybrids of 7 combinations between six Pleurotus ostreatus varieties and one Pleurotus florida variety. In this study, the results of analysis on hybridization rate, nuclear and mitochondrial DNA patterns, and colors and yields of fruit-bodies, are presented as follows. In di-mono crosses, hybrids between Pleurotus ostreatus and Pleurotus florida showed 100% of crossability as seen in those between Pleurotus ostreatus and Pleurotus ostreatus indicating that nuclei and mitochondria of a dikaryon migrated to a recipient of monokaryon. The mitochondrial DNA patterns of the hybrid strain were composed of 75% dikaryon donors and 25% monokaryon recipient. The crossability between mono-mono crossing ranges between 50 and 93.75%. 82.4% of the hybrid strain showed mitochondrial DNA patterns predominated by either parent, while the remaining 17.6% had recombinant or half-and-half combined patterns of both parents.