Objective To study the relationship between COL9A1 gene polymorphism and COL9A1 expression in the knee cartilage of Kashin-Beck disease (KBD). Methods Articular cartilage of the knee and venous blood of 65 patients with knee arthritis of KBD were collected. The whole genome DNA was extracted from the venous blood, and genotypes were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. S-P staining was used to evaluate the expression of COL9A1 in the cartilage. Results Compared to those with the TT genotype, subjects with the CC genotype were found to have a significantly higher score of articular cartilage of the knee (P <0.05). X-ray stratification study indicated that with the aggravation of the lesion, the expression of COL9A1 in the cartilage weakened, and the expression of COL9A1 in Grade II cartilage was significantly higher than that of Grade IV (P<0.05). Conclusion There is a correlation between rs6910140 in COL9A1 gene and the expression of COL9A1 in the articular cartilage of the knee in patients with KBD. The expression of COL9A1 is associated with the severity of KBD.