Abstract: This study aims to compare and analyze the characteristics, evolution, and genetic relationship of different species mitochondrial genomes（mtDNA） in Ganoderma. Three unreported Ganoderma mtDNA were assembled and annotated. The different of these mtDNA were compared combined with seven reported species in GenBank. Evolution relationship was analyzed by conserved coding protein genes and the genetic relationship was shown vis phylogenetic tree. The results showed that mtDNA of Ganoderma was a closed-loop structure of 63.616～107.774 kb, including 14 common coding protein genes, one rps3 gene, two RNA subunit genes and 23～29 tRNA genes. The genetic distance of common coding protein genes is less than 0.12 and Ka/Ks＜1, indicating that the genes were selected by purification. They were conservative relatively and did not evolve rapidly. The number of introns ranged from 11 to 39, and the number of introns in cox1 gene was the largest and great differences among species, they mainly contain RNA polymerase, GIY-YIG endonuclease or LAGLIDADG endonuclease endonuclease domain. tRNA gene exist insertion, deletion, and mutation. Most of the common coding protein genes order are highly consistent, but some Ganoderma mitochondrial genes occurred inversion and transposition. In addition, the results of genetic relationship analysis showed that Ganoderma leuccontextum had the furthest genetic relationship among the 10 Ganoderma species, which was consistent with the results of gene order analysis. In conclusion, the difference of mtDNA is mainly reflected in the number of tRNA gene and intron in Ganoderma. The genes in mitochondria are relatively conserved, G. leucocontextum has the furthest genetic relationship with other species in Ganoderma.