Mitochondrium is an important organelle in eukaryotic cells for energy conversion and ATP formation. Currently many edible fungal mitochondrial genomes have been assembled and annotated. Pleurotus pulmonarius, a member of the Pleurotaceae family in Basidiomycota, is an edible, economically important mushroom in most Asian countries. The mtDNA molecule of P.pulmonarius was found to be a single circle of 68,305 ~75,709 bp. It contained 14 core protein-coding genes, two ribosomal RNA (rRNA) subunit genes, and some transfer RNA (tRNA) genes. Introns mainly distributed in cytochrome c oxidase subunit 1 (cox1) with conservative domain of LAGLIDADG, ribosomal protein S3, and DNA polymerase II. The differential fragment sequences among strains were mainly located in the introns and intergenic regions. Results showed that the mtDNA inheritance pattern was uniparental and closely related to dikaryotic hyphal location with respect to the parent. Results also show that mtDNA inheritance is influenced by both the parental nuclear genome and mitogenome in the zone of contact between two compatible parents. During the growth, nuclear genes played a greater role than mitochondrial genes in mycelial morphology and growth rate. Both nuclear genes and mitochondrial genes could affect the nutrient content of P.pulmonarius. Mitochondrial and nuclear genes of P.pulmonarius could interact each other and act on life activities jointly.