This study investigated the effects of various culture treatments on di-n-butyl phthalate (DBP) degradation and the survival conditions of DBP-degrading bacterial strains in a soil microcosm. In the previous study, a DBP-degrading strain was isolated from activated sludge and identified by 16S rRNA as Deinococcus radiodurans. In this study, we added D. radiodurans into a soil microcosm and analyzed the structure of the whole bacterial community of the soil using a polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) technique. Meanwhile, the optimal conditions for DBP degradation were assessed by varying the temperature and initial pH of the culture, and by adding yeast extract and surfactants. The results show that the optimal conditions for DBP degradation in soil are a temperature of 35°C, a pH of 7, and the addition of Triton X-100 and yeast extract. Furthermore, the addition of D. radiodurans can also enhance DBP degradation in soil. The PCR-DGGE analysis showed that D. radiodurans could survive in the soil microcosm through 24 days of incubation. We hope that these findings may provide some useful information about the remediation of DBP in our environment.