Colorectal cancer (CRC) is the third most common cancer worldwide, affecting over 12,000 individuals in Australia each year. While CRC is often detected at a stage where resection of the primary tumour is possible, approximately 50% will relapse and die from metastatic disease. Current practice to determine clinical management is determined by the extent of cancer spread at diagnosis, tumour depth (T) and lymph node stage (N). Adjuvant 5-flurorouricil (5-FU) based chemotherapy is offered to the majority of patients with stage III CRC, reducing recurrence by approximately 40% in stage III and a subset of high risk stage II CRC patients. This approach is suboptimal, resulting in the under treatment of stage II patients who relapse (approximately 20%). Contrarily, a large number of stage III patients are over treated with only 15% responding to 5-FU as a first line treatment. There is an urgent need to identify biomarkers of prognosis and predictive benefit from 5-FU treatment for stage II and III CRC to refine outcome prediction and use of current adjuvant therapy. We aim evaluate a panel of 113 candidate CRC genes identified as exhibiting significant mutation in whole exome sequencing studies, and 74 CRC risk variants discovered in genome-wide association (GWA) studies, in 250 patients with stage II and III CRC. Here, we will present the design of our custom cancer gene panel to be evaluated, data validating the assay for use on fresh frozen as well as formalin-fixed paraffin-embedded tissue specimens and initial results.