Biochar and ryegrass have been used in the remediation of polycyclic aromatic hydrocarbons (PAHs)-contaminated soils; however, the effects of different biochar application levels on the dissipation of PAHs, bacterial communities, and PAH-ring hydroxylating dioxygenase (PAH-RHDα) genes in rhizosphere soil remain unclear. In this study, enzyme activity tests, real-time quantitative polymerase chain reaction (PCR), and high-throughput sequencing were performed to investigate the effects of different proportions of rape straw biochar (1%, 2%, and 4% (w/w)) on the degradation of PAHs, as well as the associated changes in the soil bacterial community and PAH-RHDα gene expression. The results revealed that biochar enhanced the rhizoremediation of PAH-contaminated soil and that 2% biochar-treated rhizosphere soil was the most effective in removing PAHs. Furthermore, urease activity, abundance and activity of total bacteria, and PAH-degrading bacteria were enhanced in soil that was amended with biochar and ryegrass. Additionally, the activity of 16S rDNA and PAH-RHDα gram-negative (GN) genes increased with increasing biochar dosage and had a positive correlation with the removal of PAHs. Biochar changed the rhizosphere soil bacterial composition and α-diversity, and promoted the growth of Pseudomonas and Zeaxanthinibacter. In addition, the relative abundance of Pseudomonas was positively correlated with PAH removal. These findings imply that rape straw biochar can enhance the rhizoremediation of PAH-contaminated soil by changing soil bacterial communities and stimulating the expression of PAH-RHDα GN genes. The 2% of rape straw biochar combined with ryegrass would be an effective method to remediate the PAH-contaminated soil.