Background and Objectives: Chronic Kidney Disease of uncertain or non-traditional etiology (CKDnT) is a form of chronic kidney disease of undetermined etiology (CKDu) and is not associated with traditional risk factors. The aim of this study was to investigate the association of polymorphisms rs2070744, 4b/a and rs1799983 of theNOS3gene with CKDnT in Mexican patients.Materials and Methods: We included 105 patients with CKDnT and 90 controls. Genotyping was performed by PCR-RFLP’s, genotypic and allelic frequencies were determined and compared between the two groups usingχ2analysis, and differences were expressed as odd ratios with 95% confidence intervals (CI). Values ofp< 0.05 were considered statistically significant.Results:Overall, 80% of patients were male. The rs1799983 polymorphism in NOS3 was found to be associated with CKDnT in the Mexican population (p= 0.006) (OR = 0.397; 95% CI, 0.192–0.817) under a dominant model. The genotype frequency was significantly different between the CKDnT and control groups (χ2= 8.298,p= 0.016).Conclusions: The results of this study indicate that there is an association between the rs2070744 polymorphism and CKDnT in the Mexican population. This polymorphism can play an important role in the pathophysiology of CKDnT whenever there is previous endothelial dysfunction.