Abstract Background: ADH1B rs1229984 and ALDH2 rs671 polymorphisms are associated with digestive tract tumors. In this study, we aimed to establish a method to detect ADH1B rs1229984 and ALDH2 rs671 based on PCR-SSP. The polymorphisms of rs1229984 and rs671 were detected in a Lianyungang Han healthy population; we further investigated the correlation between the distribution of the two SNPs in patients with gastric and esophageal cancer.Methods: We enrolled 200 healthy Han blood donors from Lianyungang Central Blood Bank in Jiangsu Province, and 50 patients with gastric cancer and 50 patients with esophageal cancer from the Second People's Hospital of Lianyungang (Tumor Hospital). Mismatched bases were introduced near the 3' end of the primer to increase PCR specificity. Polymorphisms of the two SNPs were detected by PCR-SSP. Standard plasmids containing SNP sites were synthesized by TA cloning. After fragment insertion was verified by sequencing, the limit of detections and detection specificities of PCR-SSP were studied. Finally, 10 samples were selected for sequencing. Data analysis was conducted with SPSS.Results: PCR-SSP was successful in the genotyping of ADH1B rs1229984 and ALDH2 rs671. Gene sequencing demonstrated an accuracy of 100%. The limits of detection of rs1229984 AA and GG were 102 and 105 copies/μL, respectively, while the LODs of rs671 AA and GG were 103 and 106 copies/μL, respectively. The specificities of rs1229984- A-R, rs1229984-G-R, and rs671-A-R were 100%, while rs671-G-R was very weakly interfered by alleles. The frequencies of rs1229984 AA, AG, and GG genotypes in healthy subjects were 42.5%, 53.5%, and 4%, respectively, and the frequencies of rs671 AA, AG, and GG genotypes were 1%, 26.5%, and 72.5%, respectively. Genotype analysis of these SNPs revealed that GG of rs1229984 is a risk genotype (p = 0.02) for esophageal cancer. Significant association was also observed between rs671 AA, AG, and the incidence of gastric cancer (p = 0.003, 0.016, respectively). Moreover, the gene frequencies of rs671AG were also statistically significant between esophageal cancer patients and healthy subjects (p = 0.000).Conclusion: We developed a method for the detection of rs1229984 and rs671 based on PCR-SSP. The sensitivity and specificity of this method were high, and operation of this method is simple and economical. We also identified the distribution of the gene polymorphisms in the Han population in the Lianyungang area, and found there was a significant difference in the gene distribution composition ratio between gastric cancer and esophageal cancer and the healthy control group.