To investigate how sodium selenite (Na<sub>2</sub>SeO<sub>3</sub>) regulated the content of vitamin C (Vc) in strawberry fruit, we explored the effects of Na<sub>2</sub>SeO<sub>3</sub> on the enzymes responsible for Vc metabolism. The findings showed that<br /> 10 mg/L Na<sub>2</sub>SeO<sub>3</sub> improved the activities of ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), glutathione reductase (GR), l-galactono-1,4-lactone dehydrogenase (GalLDH) at periods of young fruit (YFP), small fruit (SFP), middle fruit (MFP), large fruit (LFP), white fruit (WFP), colour-changed fruit (CFP) and ripen fruit (RFP). 30 mg/L Na<sub>2</sub>SeO<sub>3</sub> improved the activities of APX, MDHAR, GR and GaILDH at YFP, LFP, WFP, CFP and RFP. 60 mg/L Na<sub>2</sub>SeO<sub>3</sub> improved the activities of MDHAR, GR and GaILDH at all periods studied. In addition, 10 mg/L Na<sub>2</sub>SeO<sub>3</sub> decreased the activity of ascorbate oxidase (AAO) at WFP and CFP. 30 mg/L Na<sub>2</sub>SeO<sub>3</sub> decreased AAO activity at MFP, LFP, WFP and CFP. 60 mg/L Na<sub>2</sub>SeO<sub>3</sub> decreased AAO activity at YFP, SFP, MFP, LFP, WFP and CFP. Meanwhile, all concentrations of Na<sub>2</sub>SeO<sub>3</sub> significantly increased the contents of Vc and Se. Among different concentrations, 30 mg/L Na<sub>2</sub>SeO<sub>3</sub> had better effects on the enzymes responsible for Vc metabolism, which further improved Vc content in strawberry fruit. Besides, all concentrations of Na<sub>2</sub>SeO<sub>3</sub> increased fruit average weight, number of fruits per plant and fruit yield, compared with control. The above results indicated that Na<sub>2</sub>SeO<sub>3</sub> could improve the content of Vc in fruit and fruit yield of strawberries, especially for 30 mg/L Na<sub>2</sub>SeO<sub>3</sub>.