Background <p>Known for their unique biodiversity, the Qinling Mountains are considered the only area in which alpine biomes occur in central China. Given that the alpine biomes are particularly sensitive to global warming, understanding how alpine plants respond to climatic fluctuations is essential for the evolution and conservation of biodiversity. To address this issue, three alpine species of the Gentianaceae (<i>Gentiana crassuloides</i>, <i>G. hexaphylla</i> and <i>Swertia bifolia</i>) that represent different life types and diverse genera were selected.</p> Results <p>Genetic clustering analysis according to around 33,317 to 185,133 SNPs showed that the Qinling population was a separate lineage within each species. A high level of genetic differentiation was observed among the Qinling populations and the other populations of each species. Divergence time estimation based on plastomes and approximate Bayesian computation based on genomic SNPs showed that Qinling populations of the three Gentianaceae species originated at different periods under various patterns including primary source and hybridization. Significant signals of isolation by distance and isolation by environment were found in all three species. The redundancy and gradient forest analyses revealed that several temperature- and precipitation-related variables mainly contributed to shaping the genetic differentiation among the Qinling populations and others, indicating that the three species exhibited a similar pattern of adaptations to local environments.</p> Conclusions <p>This study unveiled the unique genetic and evolutionary features of the Qinling populations of these three species and elucidated the contributing role of both the environmental gradient and geographical isolation in genetic differentiation, which scientifically supports future conservation efforts.</p>