Two experiments were conducted to investigate the effects of supplementing a corn-soybean layer diet with either phytase, 1,25-dihydroxycholecalciferol [1,25-(OH)2D3], or their combination. The basal diet was formulated to contain 3.00% Ca and 0.33% total P. In Experiment 1, 160, 56-wk-old laying hens were randomly assigned to treatment groups fed either the basal diet alone or diets supplemented with either 600 phytase units (FTU) per kilogram feed, 5 microg 1,25-(OH)2D3/kg feed, or their combination for an experimental period of 9 wk. Experiment 2 had the same design and treatment groups except that laying hens 24 wk of age were used for 8 wk. In both experiments, phytase had a positive effect on BW and increased plasma dialyzable P, tibia bone ash, and phytate P retention. In the first experiment, the addition of phytase, 1,25-(OH)2D3, or their combination prevented a rapid decrease in egg production due to a Mycoplasma gallisepticum infection observed in hens fed the basal diet. However, no benefit in egg production was obtained in the second experiment. No effects on egg weight and egg specific gravity were observed in both experiments. These results clearly indicate that phytase, and to a lesser extent 1,25-(OH)2D3, can be used to increase the utilization of phytate P by laying hens.