Under global climate change, alfalfa (Medicago sativa L.) development is facing a large challenge of drought, which made it crucial and necessary to improve its drought tolerance through biotechnological approaches. In alfalfa, the MsSIK1 gene was identified to be one of the water use efficiency (WUE)-related genes due to its strong expression in response to dehydration. In this study, after the 26-day-long drought treatment, transgenic Arabidopsis seedlings over-expressing the MsSIK1 gene were measured to be taller than the wild-type (WT) and mutants of er-105/MsSIK1 and loss-of-function er-105 which has reduced drought tolerance capacity and serves as a WUE negative control. The length of inflorescence for the Ox MsSIK1 plants was measured to range from 192 to 213 mm, which was different among different transgenic lines; meanwhile that for the WT plants and those in er-105/MsSIK1 lines were measured to be 100 mm and the mutant line averaged 60 mm. Compared to WT and er-105/MsSIK1 and er-105 mutant lines, transgenic Arabidopsis had higher rate of long-term CO2 assimilation but lower rate of transpiration, therefore had higher instantaneous WUE, long-term WUE, and endogenous GA content. In conclusion, the MsSIK1 gene isolated from alfalfa confers an improved WUE in transgenic Arabidopsis. These results fully illustrated that the alfalfa MsSIK1 gene could improve WUE in transgenic Arabidopsis. Also they would supply genetic evidence for improving biotechnological approach for alfalfa to adapt drought stress driven by probably future climate change.