Objective To explore the relationship among the polymorphisms of CYP2R1 gene at rs10741657, rs10766197, and rs1993116 loci, vitamin D levels and susceptibility to tuberculosis. Methods 174 tuberculosis patients from a hospital in Qinghai Province were selected as the case group and 202 healthy individuals underwent physical examination during the same period were selected as the control group. Polymerase chain reaction (PCR) was used to amplify the rs10741657, rs10766197 and rs1993116 loci, and determine the genotypes. The vitamin D level was determined by enzyme-linked immunosorbent assay (ELISA). Results rs10741657 gene polymorphism was associated with tuberculosis(χ²=6.884, P=0.032), while no association was found between rs10766197, rs1993116 polymorphisms and tuberculosis. Among tuberculosis patients, individuals carrying the GG genotype at the rs10741657 locus had lower vitamin D level than those with the AA genotype (P=0.033) and AG genotype (P=0.034). The genotypes at the rs10766197 and rs1993116 loci were not found to be associated with vitamin D levels. Conclusion Gene polymorphism at the rs10741657 locus is associated with vitamin D levels and the development of tuberculosis.