Abstract: The Calvin cycle is an irreplaceable pathway for photosynthetic organisms to assimilate CO2 from the air and therefore plays a crucial role in plant growth and development. The ribulose-phosphate 3-epimerase (RPEase: EC 5.1.3.1) is one of the integral enzymes in the Calvin cycle regeneration phase. In our study PdRPE gene was cloned from fast-growing hybrid poplar NE-19 (Populus nigra × (Populus deltoids × Populus nigra)), and then PdRPE was transformed into Arabidopsis thaliana by the floral dip method. Using PCR and Green Fluorescent Protein (GFP) detection methods, we obtained the CaMV35S:PdRPE:GFP transgenic plants. Then we validated the growth status and physiological indexes of wild type (Col-0), overexpressing (OxPdRPE), mutant (rpe) and complementary (rpe/PdRPE) lines. The results indicated that in overexpressing lines RPEase activity increased significantly (P<0.05). When well-watered, the overexpressing lines had lower leaf stomatal density, larger stomatal size, improved water use efficiency and higher net photosynthetic rate, showing a better growth condition and more starch accumulation. When exposed to 10 days of short-term drought, the net photosynthetic rate and water use efficiency of overexpressing lines were still significantly higher than those of the other three strains (P<0.01). Therefore, we reach the conclusion that over-expression of RPEase gene can promote the plant growth and development and enhance the resistance of short-duration drought in Arabidopsis thaliana.