In this study, the major factors affecting both cell growth and astaxanthin production of Chlorella zofingiensis under dark were identified and optimized through statistical methods. A Plackett-Burman design was initially used to evaluate seven culture factors, of which glucose, $NaNO_3$, and $MgSO_4{cdot}7H_2O$ were found to be the most important in affecting C. zofingiensis. The central composite design and response surface methodology (RSM) were further used to optimize these factors. The optimized glucose, $NaNO_3$, and $MgSO_4{cdot}7H_2O$ concentrations for cell growth were 46.7, 1.13, and 0.125 g/L $MgSO_4{cdot}7H_2O$ respectively, and for astaxanthin production were 35.2, 0.281, and 0.023 g/L, respectively. After the optimization, a two-stage culture strategy was employed to further maximize astaxanthin production, and the results showed that the astaxanthin yield of C. zofingiensis could reach 15.1 mg/L, which was 74% higher than that achieved in batch culture using the basal medium (i.e., 8.7 mg/L).