In order to explore the effect of adding Bacillus subtilis on the microbial community structure, nutrition of bioflocs and water quality in the process of system construction, the biofloc technology-aquaculture system was completed, the additive concentration of B. subtilis was group A, 10⁶ CFU/mL, group B,10⁵ CFU/mL, group C, 10⁴ CFU/mL, group D, 0 CFU/mL. The experimental results showed that:The concentration of B. subtilis had significant effect on the concentration of TSS and FV-30; The concentration of B. subtilis had a significant effect on the amino acid content and C/N of the floc; The concentration of B. subtilis had no significant effect on crude lipid and ash content; The amino acid and crude protein contents of adding B. subtilis groups were higher than those of the non-added group; The additive of B. subtilis had a significant effect on the relative abundance at phylum and genus level of bacteria; and increasing the concentration of B. subtilis additive led to an increase in the concentration of Bacillus in the biofloc. B. subtilis can be effectively enriched on bioflocs. The addition of B. subtilis can effectively improve the nutrient content of bioflocs, which is conducive to providing protein source for aquaculture.