Abstract: Understanding of the photosynthetic and physiological heterogeneities of seedling provenances in response to climatic variations across different habitats provides critical guidance for the scientific introduction of forestry. In this study, one-year-old seedlings of Thuja plicata from three provenances (BC, Pur, Py) were transplanted in southern (Nanchang, Jiangxi) and northern (Tongzhou, Beijing) experimental nurseries, aiming to illustrate the differences in seedling growth, photosynthetic and physiological traits, and the influences of climatic differences between plantation and provenance sites on seedling photosynthetic performance were also investigated. Results show that: (1) The relative growth rate of seedlings was significantly higher in southern site than that in northern site, suggesting that habitats posted a significant influence on the growth of seedlings. In southern habitat, Pur exhibited superior growth performance when compared to other provenances. (2) Photosynthetic parameters were significantly influenced by provenances, habitats, light intensity, and their interactions. In northern habitat, BC demonstrated a higher net photosynthetic rate (P_n), stomatal conductance (G_s), intercellular CO₂ concentration (C_i), and transpiration rate (T_r), while Py had the highest water use efficiency (WUE). As a contrast, Py showed a better performance in G_s, C_i, and T_r, with the lowest WUE in southern habitat; Pur displayed relatively consistent performance across both habitats. (3) In northern habitat, Py had significantly higher maximum photochemical efficiency and actual photochemical efficiency than Pur; the photochemical quenching of Py was significantly higher in southern habitat than that in northern habitat. (4) Seedling photosynthetic parameters were significantly correlated with the climatic distance between plantation and provenance sites. P_n was only negatively regulated by the minimum temperature difference (ΔT_min). Additionally, soil moisture difference and ΔT_min had the highest explanatory contribution for variations in G_s and WUE, promoting G_s and inhibiting WUE, respectively. In conclusion, the growth and photosynthetic traits of Thuja plicata seedlings exhibit significant provenance and habitat differences, indicating that both provenance selection and habitat matching should be considered in introduction and cultivation to enhance tree adaptability after transplantation.