Brain-derived neurotrophic factor (BDNF) Val66Met polymorphism is reported to be associated with cognitive dysfunction, an important comorbidity factor in patients with type 2 diabetes mellitus (T2DM), especially in elderly populations, however, the underlying pathophysiological mechanisms are unclear. This study was performed to investigate the association between BDNF Val66Met polymorphism and mild cognitive impairment (MCI) in elderly patients with T2DM. In total, 105 MCI and 105 normal cognition controls of T2DM patients were enrolled; all of the patients underwent neuropsychological assessments. BDNF Val66Met polymorphism was genotyped via TaqMan SNP genotyping assay. Data from clinical and laboratory-based examinations were collected. The frequency of the BDNF Met allele was significantly higher in the MCI group than in the controls. Multiple regression analysis indicated an association of the Met allele with MCI in patients with T2DM (OR = 2.54; 95% CI 1.33-4.84; p = 0.005). Stratified by educational level, the BDNF Met allele was significantly associated with MCI in elderly T2DM patients (OR = 3.29; 95% CI 1.26-8.57; p = 0.015) among the group of low educational levels (< 12years); however, the association was insignificant among those with higher educational levels. BDNF Met allele carriers showed a higher frequency of MCI than Val/Val homozygotes in elderly T2DM patients. However, this association was only significant in patients with low education levels. BDNF Val66Met polymorphism may have a potential role in MCI in elderly T2DM patients, especially those with low educational levels.