Abstract: To determine the optimum carbon source and dosage for bio-floc formation in Penaeus monodon culture system, we added three kinds of carbon source including glucose, sucrose and molasses in the culture system at C/N ratio of 20 : 1. The sucrose was added into the culture system at daily feeding ratios of 0%, 25%, 50%, 75% and 100%. Results show that all carbon sources tested in the present study could significantly increase the special growth rate (SGR) of P.monodon (P < 0.05). Sucrose could significantly increase the survival rate (SR) of P.monodon (P < 0.05). Compared with the other groups, 75% of sucrose could significantly improve the SGR and SR of P.monodon (P < 0.05), and sucrose could promote heterotrophic bacteria and phytoplankton reproduction, and indirectly reduce concentration of harmful substances in water such as NH₄-N and NO₂-N, but adding excessive sucrose would be counterproductive. The results reveal that adding 75% of sucrose was the most suitable condition for bio-floc formation in P.monodon culture system, which could promote the growth of P.monodon.