Gastroesophageal reflux disease (GERD) may lead to Barrett's esophagus (BE). Previously, a large genome-wide association study found two germline markers to be associated with BE, FOXF1 rs9936833 (C allele) and MHC rs9257809 (A allele). This study evaluated whether these two polymorphisms are associated with gastroesphageal acid reflux as measured by 24-hour pH testing. Patients with acid reflux symptoms referred for esophageal manometry and 24-hour pH monitoring at University Health Network (Toronto, ON) were enrolled. DNA extracted from blood was genotyped using a Taqman Polymerase Chain Reaction (PCR) assay. DeMeester scores of ≥14.7 or prior evidence of reflux esophagitis on endoscopy defined individuals with esophageal acid reflux. Logistic regression analysis, adjusted for clinical risk factors, was used to calculate odds ratios with 95% confidence intervals for each polymorphism in relation to the presence of acid reflux. Of 182 patients, the median age was 50 years and 62% were female; 95 (52%) met the definition of GERD. In the multivariable analysis, both FOXF1 rs9936833 (OR = 1.82; 95%CI: 1.12–2.96; P = 0.02) and MHC rs9257809 (OR = 9.36; 95%CI: 2.92–29.99; P < 0.001) remained significantly associated with presence of acid reflux. When both polymorphisms were placed in the same model, the adjusted ORs were 2.10 (95%CI: 1.24–3.53; P = 0.005) and 10.95 (95%CI: 3.32–36.09; P < 0.001), respectively. The association for risk allele C in FOXF1 rs9936833 and risk allele A in MHC rs9257809 with the presence of acid reflux suggests a potential pathophysiologic mechanism for the role of genetic influences in BE development.