Objective To explore the conditions of transfecting bone morphogenetic protein 2 (BMP2) gene into human periodontal ligament fibroblasts (HPDLFs) by SonoVue and its effects on osteogenesis differentiation of HPDLFs. Methods The HPDLFs were primary cultured and transfected under different conditions (concentrations of SonoVue, different ultrasound exposure intensity and time) to screening for the optimum parameters. Cells were randomly divided into four groups:SonoVue+plasmid+ultrasound group, liposome+plasmid group, ultrasound group, and blank control group. After transfection with the corresponding strategies, the alkaline phosphatase activity was detected on day 3, 5, 7 and 9 d, and alizarin red staining was applied to check the calcium nodus on day 21 d, respectively. The mRNA expression of osteopontin (OPN), bone sialoprotein (BSP), typeⅠcollagen (ColⅠ), osteocalcin (OCN), and runt-related transcription factor-2 (Runx2) was detected by RT-PCR on day 7, 10 and 14 d of transfection. Results The transfection rate and the cell survival rate were relatively high under the following condition:0.8 W/cm², 90 s and 20% SonoVue solution, and they were selected as the final transfection parameters. Both alkaline phosphatase activity and matrix mineralization content in SonoVue+plasmid+ultrasound group were significantly higher than those in blank control group and ultrasound group (P<0.05) at all time points. RT-PCR results revealed that the expression of OPN, BSP, ColⅠ, OCN and Runx2 mRNA in SonoVue+plasmid+ultrasound group was significantly higher than that in the ultrasound group and blank control group (P<0.05), but was similar to that in the liposome+plasmid group (P>0.05). Conclusion The BMP2 gene can be successfully transfected into HPDLFs by SonoVue and can induce osteogenesis differentiation of HPDLFs, which lays a foundation for its application in periodontal tissue engineering.