The production of bacterial cellulose (BC) from Gluconacetobacter xylinus could be improved by the two-stage cultivation strategy. The jar fermentor was applied at first stage to increase cell concentration. At the second stage, the bacteria statically grew within beaker. Three different fraction of cell volumes were cultured in modified YPD-glucose medium or YPD-molasses one. In the modified YPD-molasses medium, the BC yield were highest in this study when the consuming rate of reducing sugars (RS) was 0.181 g-dry BC L-1 d-1 and the BC production rate was 0.183 g-dry BC/g-RS, respectively. The modified YPD-glucose medium could get the maximum pellicle thickness of 5.56±0.64 mm and water content of 99.4%, respectively.