To explore the association of polymorphisms in the region of three neighboring genes TRIT1, MYCL1 and MFSD2A with risk and clinicopathological features of gastric cancer, 19 tagging SNPs in this region were genotyped using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry in a case-control study of 610 Chinese gastric cancer patients and 608 cancer-free controls. MFSD2A rs4233508 T>C CC genotype was associated with an increased risk of gastric cancer in younger patients and an increased risk of moderately/well-differentiated intestinal-type gastric cancer (adjusted odds ratio [OR], 1.74 and 1.50, respectively). TRIT1 rs11581557 T>G TG was associated with lymph node metastasis (TG versus TT/GG, adjusted OR, 1.64). MFSD2A rs12083239 GC genotype and TRIT1 rs2172362 or rs230310 homozygous genotype were associated with Lauren's classification (GC versus GG, adjusted OR, 1.69; GC versus GG/CC, adjusted OR, 1.74) and tumor site (rs2172362: CC versus CT, adjusted OR, 1.71; CC/TT versus CT, adjusted OR, 1.62; rs230310: CC versus CT, adjusted OR, 1.75; CC/TT versus CT, adjusted OR, 1.67) of gastric cancer, respectively. One TRIT1 haplotype, CCGT, was associated with lymph node metastasis and tumor site of gastric cancer (CCGT versus TTTT, adjusted OR, 1.91 and 1.55). This is believed to be the first report that several tagging SNPs and haplotypes in TRIT1, MYCL1 and MFSD2A region are significantly associated with risk and clinicopathological features of gastric cancer in a Chinese population. The findings might be useful for risk assessment and prognosis prediction of gastric cancer.