Background: Recent data show that patients with left sided colon tumors (LT) have better survival and respond differently to biologics compared to patients with right-sided tumors (RT), likely due to molecular differences. We sought to examine these differences. Methods: Primary colorectal tumors (n = 1730) with origins clearly defined as RT (cecum to hepatic flexure; n = 273), LT (splenic flexure to sigmoid colon; n = 585), or rectal (RC; n = 872) were examined by NextGen sequencing, protein expression and gene amplification. Tumor mutational load (TML) was calculated in 1001 of these tumors using only somatic nonsynonymous missense mutations. Chi-square was used for comparison. Results: When compared to LT, RT carried a significantly higher rate of BRAF (25% vs 7%; p < 0.0001), PTEN (5.4% vs 1.3%; p = 0.008), and ATM (4% vs 1%; p = 0.04) mutations. RT were likely to have more MSI-high tumors (22% vs 5%; p < 0.0001) and PD-1 overexpression (58% vs 44%; p = 0.01). There were no differences in the rate of KRAS (50% vs 42%; p = 0.07) or NRAS mutations (2.2% vs 3.4%; p = 0.4). When compared to RC, RT had a higher rate of BRAF (25% vs 3%; p = 7E^-07), PIK3CA (22% vs 11%; p = 0.001), CTNNB1 (3% vs 0.3%; p = 0.02); ATM (3% vs 1%; p = 0.04), PTEN (5% vs 1%; p = 0.004), and BRCA1 mutations (4% vs 0%; p = 0.02), and a lower rate of TP53 (56% vs 71%; p = 0.001) and APC (53% vs 66%; p = 0.003) mutations. When compared to RC, LT showed higher rates of BRAF (6.7% vs 3.2%; p = 0.04) and CTNNB1 (2.1% vs 0.3%; p = 0.04) mutations, and a higher rate of MSI-high tumors (4.6% vs 0.7%; p = 0.04), whereas RC had a higher rate of KRAS mutation (50% vs 42%; p = 0.04). There were no differences between RT, LT, and RC for the frequency of PD-L1 (2%, 2%, and 1%) or Her-2 (1%, 2%, and 3%) overexpression, although Her-2 amplification was significantly different (1%, 3%, and 5%, RT vs RC; p = 0.03). Mean TML was 12, 11, and 8 mutations/megabase for RT, LT, and RC, respectively (RT vs RC; p = 0.01). There was a correlation between TML and PD-L1 (p = 0.04) and PD-1 (p = 0.01). Conclusions: Tumors arising in the right colon carry genetic alterations that are different from LT as well as RC. However, it appears that CRCs carry a continuum of molecular alterations from the right to the left side, rather than displaying sharp, clear-cut differences.