In this study, a lab-scale sequencing batch bioreactor (SBR) with hydraulic retention time (HRT) of 8 h was established to treat 50 mg/L 4-chlorophenol (4-CP), and the temporal responses of 4-CP on the performance and the expression level of related functional genes in sludge were explored. Results showed that after the SBR was acclimated with the cometabolic carbon sources of 4-CP and methanol, the pollutants were effectively degraded. The pollutants removal was influenced by the polysaccharides and proteins in sludge. The real-time PCR showed that except for the gene p450, the prompt removal of pollutants before 2 h caused the significant upregulation of the other functional genes from the genus Pseudomonas in sludge, while the downregulation occurred from 2 to 8 h with the progressive removal of pollutants in SBR cycle, indicating that the removal of pollutants was closely related to the functional genes expression.