Tomato (Solanum lycopersicum cv. ‘Moneymaker’) was transformed with the choline oxidase gene codA from Arthrobacter globiformis, which was modified to allow for targeting to both chloroplasts and the cytosol. Glycine betaine (GB) was accumulated in transformed plants, while no detectable GB was found in wild-type (WT) plants. Compared to WT plants, transgenic lines showed significantly higher photosynthetic rates (Pn) and antioxidant enzyme activities and lower reactive oxygen species (ROS) accumulation in the leaves when exposed to salt stress. Furthermore, compared with WT plants, K⁺ efflux decreased and Na⁺ efflux increased in roots of transgenic plants under salt stress; resulted in lower Na⁺/K⁺ ratios in transgenic lines. The exogenous application of GB also significantly reduced NaCl-induced K⁺ efflux and increased Na⁺ efflux in WT plants. A qRT-PCR assay indicated that GB enhanced NaCl-induced expression of genes encoding the K⁺ transporter, Na⁺/H⁺ antiporter, and H⁺-ATPase. These results suggest that the enhanced salt tolerance conferred by codA in transgenic tomato plants might be due to the regulation of ion channel and transporters by GB, which would allow high potassium levels and low sodium levels to be maintained in transgenic plants under salt stress condition.