The conventional treatment of dark coloured textile wastewater using chemical coagulation generates large volume of sludge, which requires further treatment and disposal. In the present investigation, a systematic optimization study of the important variables influencing the decolorization of Reactive Orange-16 (RO-16) and Reactive Red-35 (RR-35) dyes by the white-rot fungus (Trametes versicolor) was carried out. A full factorial central composite design was employed for experimental design and optimization of results. The effect of concentrations of dye, glucose and ammonium chloride on decolorization was studied and optimized using Response Surface Methodology (RSM). Maximum decolorization of 94.5% and 90.7% for RO-16 and RR-35 was obtained at optimum concentrations of dye, glucose and ammonium chloride i.e., 0.66, 17.50 and 2.69 g/L for RO-16 and 0.68, 16.67 and 2.13 g/L for RR-35, respectively.