A mitochondrial (mt) transcriptional unit, nad3-orf156, was studied in the nucleus-cytoplasm hybrids of wheat with D/D2 plasmons from Aegilops species and their parental lines. A comparative RFLP analysis and sequencing of the random PCR clones revealed the presence of seven sequence types and their polymorphic sites were mapped. All the hybrids possessed the paternal copies besides the maternal copies. More paternal copies were present in the D2 plasmon hybrids, whereas more maternal copies were present in the D plasmon hybrids. Two major copies were present with different stoichiometries in the maternal Aegilops parents. However, only a major D plasmon copy was detected in the hybrids, irrespective of their plasmon types. The hexaploid wheat parent (AABBDD genome) possessed the major D plasmon copy in approximately 5% stoichiometry, while no D plasmon-homologous copies were detected in the tetraploid wheat parent (AABB genome). The results suggest that the observed mtDNA heteroplasmy is due to paternal contribution of mtDNA. The different copy stoichiometry suggests differential amplification of the heteroplasmic copies among the hybrids and the parental lines. All editing sites and their editing frequencies were conserved among the lines, and only the maternal pattern of editing occurred in the hybrids.