To investigate the relativity of MGMT(O-6-methylguanine-DNA methyltransferase) gene methylation from patients with protein expression and osteosarcoma necrosis rate after chemotherapy. Fifty-one oteosarcoma tissues were collected, Methylation of MGMT gene promoter was detected by methylation-specific PCR method, and protein expression of MGMT was examined by immunohistochemistry procedure, the relationship between methylated MGMT gene expression and patients response to chemotherapy was analyzed. The positive ratio of methylation MGMT gene promoter in 51 patients was 23.5% (12 in 51). Negative percentage of protein expression of MGMT was 27.5% (14 in 51). It seemed that methylation of MGMT gene in osteosarcoma tissues had no evident relationship with the patient's age, sexuality, and the size and type of neoplasms, etc. The necrosis rates of methylated MGMT of osteosarcoma (tumor grade from I to IV) were 0 (0/51), 3.9% (2/51), 5.9% (3/51), 13.7% (7/51), respectively. In contrast, the necrosis rates of unmethylated MGMT of osteosarcoma (tumor grade from I to IV) were 45.1% (23/51), 25.5% (13/51), 3.9% (2/51), 2.0% (1/51), respectively. It suggest that methylated and unmethylated MGMT gene of osteosarcoma have significant difference in protein expression. The unmethylated MGMT gene has higher positive protein expression (u = -4.92, P < 0.001). Methylation of MGMT gene has higher tumor necrosis rate in osteosarcoma patients. Methylation in MGMT promoter may be important for judging the effect of chemotherapy in Osteosarcoma patients.