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AIM: Recombined plasmid pETNF-P16 was constructed toinvestigate its expression properties in esophagealsquamous carcinoma cell line EC9706 induced by X-rayirradiation and the feasibility of gene-radiotherapy foresophageal carcinoma.METHODS: Recombined plasmid pETNF-P16 was constructed and transfected into EC9706 cells with lipofectamine. ELISA,West blot, and immunocytochemistry were performed to determine the expression properties of pETNF-P16 in EC9706 after transfection induced by X-ray irradiation.RESULTS: Eukaryotic expression vector pETNF-P16 was successfully constructed and transfected into EC9706 cells.TNFα expressions were significantly increased in the transfected cells after different doses of X-ray irradiation than in those after 0Gy irradiation (1 192.330-2 026.518 pg/mL,P＜0.05-0.01), and the TNFα expressions and P16 were significantly higher 6-48 h after 2 Gy X-ray irradiation (358.963-585.571 pg/mL, P＜0.05-0.001). No P16 expression was detected in normal EC9706 cells. However, there was strong expression in the transfected and irradiation groups.CONCLUSION: X-ray irradiation induction could significantly enhance TNFα and P16 expression in EC9706 cells transfected with pETNF-P16 plasmid. These results may provide important experimental data and therapeutic potential for gene-radiotherapy of esophageal carcinoma.