Background: Among patients with diabetic polyneuropathy, the status of folic acid, homocysteine, and nerve conduction studies (NCS) variations has been associated with methylenetetrahydrofolate reductase (MTHFR) gene polymorphisms. The current study was aimed to assess the effects of methylenetetrahydrofolate reductase (MTHFR) gene polymorphism in development of Diabetic Peripheral Neuropathy. Subjects and Methods: This case control study was carried out in outpatient clinic of endocrinology and Medical Biochemistry department, Zagazig University Hospital during a period from February to December 2018. The study included (129) participants were classified into three groups: Group (1): Included 43 apparently healthy peoples who served as a control group. Group (2): Included 43 patients diagnosed with type 2 DM according ADA criteria for diabetes. Group (3): Included 43 diabetic patients with diabetic peripheral neuropathy according to the standard Neuropathy Symptom Score (NSS) and Neuropathy Disability Score (NDS) criteria for diabetic neuropathy diagnosis. Results: the frequency of C677T MTHFR gene polymorphism was significantly higher in the patients with DPN compared with the patients without neuropathy. Conclusion: MTHFR gene polymorphism was associated with increased risk for DPN development.