Abstract: Objective To analyze the molecular characteristics of drug-resistance associated genes of multi-drug resistant Mycobacterium tuberculosis (MDR-TB). Methods Drug-resistant associated genes of inhA, katG, oxyR-ahpC (isoniazid), rpoB (rifampicin), gyrA (ofloxacin) and rrs (kanamycin) of 138 MDR-TB strains and 45 sensitive strains isolated in China were detected. Results Among 138 MDR-TB strains, 14.4% had mutations in inhA, 72.5% had mutations in katG, and 15.9% had mutations in oxyR-ahpC. The detection rate of the drug resistant mutations to isoniazid of the 3 gens mentioned above was 90.6%. 94.2% had rpoB mutation, 74.5% had gyrA mutation, and 61.1% had rrs mutation. The majority of mutations were at katG315 (66.7%), inhA-15 (9.4%), oxyR-ahpC-10 (5.1%), rpoB516 (13.8%), 526 (26.1%), 531(49.3%), gyrA90 (21.6%), 94 (51%) and rrs1401 (61.1%). Conclusion The most common gene mutations associated with isoniazid, rifampicin, ofloxacin and kanamycin resistance in MDR-TB strains in China were at katG315, inhA-15, rpoB516, 526, 531, gyrA90, 94 and rrs1401, respectively.