To investigate the frequencies of mitochondria DNA (mtDNA) tRNA(Leu (UUR)) point mutation A3243G and NADH dehydronase subunit 1(ND1) gene point mutation G3316A in Wenzhou area of Zhejiang Province, and to explore the correlation between these mutations and the clinical manifestations in patients with type 2 mellitus diabetes(T2DM). Two hundreds and forty-four unrelated patients with T2DM and 156 healthy subjects without family history of T2DM were enrolled in Wenzhou area in this study and screened for the point mutations mentioned above with polymerase chain reaction (PCR) and restricted fragment length polymorphism(RFLP) analysis. The heterogeneous mutations were confirmed with DNA sequencing and denaturing high performance liquid chromatography (DHPLC) following T-A cloning of PCR products. The percentage of A3243G mutation in group of patients with T2DM and control were 0.410% and 0.0% (1/244 vs 0/156), respectively; however, there's not any significant difference between these two groups in frequency of A3243G mutation (P>0.05). G3316A mutation was detected in 4 of 244 cases with T2DM (1.639%) and 2 of 156 healthy controls (1.282%), showing that there's also no statistic difference between these two groups in frequency of G3316A mutation (P>0.05). It's shown that the frequency of mtDNA tRNA(Leu (UUR)) A3243G mutation is fairly low in patients with T2DM in Wenzhou area. Thus it's reasonable to assume that this mutation may not be involved in the development and progression of T2DM. Furthermore, it's demonstrated that the rate of G3316A mutation of mtDNA ND1 gene is rare in patients with T2DM in Wenzhou area and this mutation also happened in healthy control. It's suggested that G3316A mutation is just a gene polymorphism of mtDNA and not related to the pathogenesis of T2DM.