TNM staging in colon cancer has several limitations. Prognostic molecular markers are now being developed to address these limitations. The aim of this study was to identify a combination of genes and markers whose expression is predictive of nodal status and outcome in colon cancer. The expression of 12 genetic markers were examined in 66 node-positive and 65 node-negative T3 colon cancers. Gene expression was quantified using real-time polymerase chain reaction. Microsatellite instability status was available through the registry. Association with lymph node status was examined using univariate and multivariate logistic regression. Thymidylate synthase expression was statistically significantly associated with lymph node status (odds ratio 0.36; 95% confidence interval: 0.16-0.81). Microsatellite instability and the other genes were not associated with nodal status. Multiple logistic regression did not identify a significant multivariate predictive model. Decreased expression of thymidylate synthase is associated with a higher risk of lymph node metastasis in patients with T3 colon cancers. Microsatellite instability and the expression of other genes are not predictive of nodal status in this population. Thymidylate synthase gene expression may help identify patients at greater risk for progression of disease.