BACKGROUND: Poplars are important woody plants, which are widely distributed in the forests from the subtropics to the north of the Northern Hemisphere. Poplars have high ecological and economic value. However, there are frequent interspecific and intraspecific hybrids in Populus, resulting in a large number of intermediate taxa, which makes the morphological identification of Populus very challenging. Plastid genome is an important tool to study the evolutionary relationship of plants. Therefore, comparison and phylogenetic analysis were carried out based on the population chloroplast genomes of 34 individuals from 7 taxa. RESULTS: In this study, seven newly assembled and annotated chloroplast genomes of Populus were reported. They all had typical quadripartite structures with the same GC content (37.6%), but there were differences within the population, and the genome size ranged from 155,736 bp to 156,812 bp. In all Populus species, 134 genes were identified, including 88 protein coding genes (PCGs), 37 tRNA and 8 rRNA genes. The gene sequences alignment of different taxa showed that the gene sequences and content were relatively conservative, there was no gene rearrangement, and only 3 highly variable regions (psbZ-trnG, ndhC-trnV and trnN-trnR) were identified, which can be used as molecular markers. Most PCGs had high codon usage bias and 3 positive selection genes (rps7, rps12 and rpl16) have been identified. The analysis of population genetic structure and phylogeny showed that the chloroplast genomes supported that Populus was a monophyletic taxon, which could be divided into four sections (Abaso, Turanga, Populus and ATL (Aigeiros, Tacamahaca and Leucoides)). Among them, P. dafengensis, P. butuoensis and P. szechuanica had the closest genetic relationship, P. gonggaensis and P. cathayana had the closest genetic relationship, it was speculated that the taxa of Sect. Tacamahaca may be the main female parent of the three new taxa from Sect. Leucoides. CONCLUSION: In general, this study provides valuable insights for new species identification, phylogenetic relationships, breeding and resource development, and genetic diversity of Populus.