Genomics research of Populus deltoides, an important timber species that is widely planted worldwide, is an important part of poplar breeding. Currently, the nuclear and chloroplast genome of P. deltoides have been sequenced, but its mitochondrial genome (mitogenome) has not been reported. To further explore the evolution and phylogeny of P. deltoides, the mitogenome of P. deltoides I-69 was assembled using reads from Nanopore and Illumina sequencing platforms and found to consist of 802,637 bp and three circular chromosomes (336,205, 280,841, and 185,591 bp) containing 58 genes (34 protein-coding genes, 21 tRNA genes, and 3 rRNA genes). RNA analysis in combination with several species showed significantly fewer RNA editing sites in the mitogenomes of poplar and other angiosperms than in gymnosperms. Sequence transfer analysis showed extensive mitogenome rearrangements in Populus species, and with evolution from lower to higher plants, tRNA transfer from chloroplasts to mitochondria became increasingly frequent. In a phylogenetic analysis, the evolutionary status of P. deltoides was determined, and the section Populus was supported. Our results based on the first report of a multi-circular conformation of the Populus mitogenome provide a basis for further study of the evolution and genetics of P. deltoides and other Populus species and for breeding programs.