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Poplars are widely distributed in the northern hemisphere and have good adaptability to different living environments. The accumulation of genome and transcriptome data provides a chance to conduct comparative genomics and transcriptomics analyses to elucidate the evolutionary patterns of Populus phylogeny. Transcript sequences of eight Salicaceae species were downloaded from public databases. All of the pairwise orthologues were identified by comparative transcriptome analysis in these species, from which we constructed a phylogenetic tree and estimated the rate of divergence. The divergence times of the phylogenetic clades were mainly estimated during the Middle Miocene Climate Transition (MMCT) to Quaternary Ice Age. We also identified all of the fast-evolving sequences of positive selection and found some resistance genes that were related to environmental factors. Our results suggest that drought-, H2O2- and cold-stress genes are involved in positive selection along with the paleoclimate change. These data are useful in elucidating the evolutionary patterns and causes of speciation in the Populus lineage.