Human DNA polymerase β (DNA polymeraseβ (polβ)) is a small monomeric protein which is essential for short-patch base excision repair (BER). It plays an important role in regulating the radiation sensitivity of tumor cells in the course of tumor radiation therapy. In this study, qRT-PCR and Western blot assays were used to quantify polβ expression levels in esophageal carcinoma (EC) cells that were transfected with polβ small interfering RNA (siRNA). Cell counting Kit-8 (CCK-8), flow cytometry, and Hoechst/PI stain assays were conducted to evaluate the effects of silencing polβ on the radiotherapeutic sensitivity of EC cells. We found that the expression levels of polβ in EC cells were significantly decreased after transfection with polβ siRNA. Then, we found that polβ silencing increased the sensitivity of EC cells to radiation therapy. In conclusion, our study paves the way for a better understanding of the mechanism of the polβ gene in DNA repair, and we propose that RNA interference technology will have important applications in gene therapy of EC and other cancers in the future.