Objective To study the correlations between O (6) -methylguanine-DNA-methyltransferase (MGMT) gene promoter methylation status in malignant glioma tissues and both MGMT protein expression and survival prognosis in these patients, and evaluate the significance of MGMT gene methylation status analyzing with methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) method in chemotherapy of brain glioma.Methods Thirty-nine patients with gliomas confirmed by pathology (WHO grade Ⅲ and grade Ⅳ)were collected in our study; the patient's overall survival (OS) after chemotherapy was tracked.MGMT protein expression of glioma tissues was detected by immunohistochemical staining,and MGMT promoter methylation status was detected by MS-MLPA method. Results Statistical difference of OS time was noted between patients with MGMT-negative and patients with MGMT-positive/-weak-positive (P=0.003).The prognosis in patients with positive MGMT protein expression was obviously poorer than that in patients with negative expression. In the groups of MGMT promoter un-methylation, mild hypermethylation, moderate hypermethylation and extensive hypermethylation, significant statistical difference of OS time was noted between each 2 groups (P＜0.05); the higher degree of methylation,the better prognosis. Statistical correlation was noted between MGMT protein expression and promoter methylation status (r=0.697,P=0.000); the higher degree ofmethylation,the lower protein exression of MGMT. Conclusion Both MGMT protein expression and promoter methylation status can be regarded as prognostic indicator of OS in patients with malignant glioma accepted alkylating agent chemotherapy; MS-MLPA is a reliable method to detect MGMT gene promoter methylation status.