[ Objective] The paper presents the diumal changes of photosynthesis and transpiration of different alfalfa varieties and their relationship with the associated physiological and ecological factors during branching stage, so as to provide a basis for the development, utilization, and breed- ing of alfalfa. [ Method] Under natural conditions, the diurnal changes of net photosynthetic rate (Pn), transpiration rate (Tr), the relevant physio- logical factors including leaf temperature (TI), stomatal conductance (Gs) and intemal COn concentration (Ci), as well as the relevant physiologi- cal factors including photosynthetic available radiation (PAR), CO2 concentration in field (Ca) and air temperature (Ta) were measured in four al- falfa varieties (Algonguin, WL323 HQ, WL414, and Millionaire). The water use efficiency (WUE) and light use efficiency (LUE) were calculated, and the correlation among them was also analyzed. [Result] The Pn, Tr, PAR and Ta of the four varieties appeared to vary in a single-peak curve; the sequence of WUE was WL323 HQ ~ Algonguin ~ WL414 ~ Millionaire; there was no significant difference in LUE of the four alfalfa varieties; coef- ficient analysis showed that Pn was mainly affected by PAR, Gs, and Ci, while Tr by PAR and Ta. [ Conclusion] WL323 HQ is the variety with high Pn, high WUE and low Tr, and it has strong adaptability to drought. In four alfalfa varieties, PAR, Ta, Gs, and TI are the primary determining fac- tors while Ca and Ci the limiting factors of Tr; Gs is the primary determining factor while Ci the limiting factor of Pn.