Colorectal cancer (CRC) is a multifactorial disease that develops as a result of well-established genetic factors such as mutation of the tumor suppressor Adenomatous Polyposis Coli (APC) gene. However, other poorly characterized factors such as the gut microbiota (GM) likely play an important role in tumor development and progression. Our laboratory has found significant differences in both small intestinal (SI) and colonic tumor numbers between two ApcMin mouse colonies; the C57BL/6J-ApcMin (B6-Min/J) from the Jackson Laboratories, and the C57BL/6JD-ApcMin (B6-Min/D) closed colony that has been maintained at the University of Wisconsin. It is unclear whether underlying host genetic factors or differences in the GM are responsible for these phenotypic differences. To determine the potential impact of GM on disease phenotype, we used complex microbiota targeted rederivation (CMTR) to rederive isogenic embryos of the two Apc mutant colonies onto CD1 surrogate dams that possessed GM from either The Jackson Laboratory (Crl:CD1GMJAX) or Envigo (Hsd:CD1GMHSD). We generated a total of four ApcMin groups: B6-Min/JGMJAX, B6-Min/JGMHSD, B6-Min/DGMJAX, and B6-Min/DGMHSD. At three months of age, all animals were sacrificed to determine both SI and colonic tumor multiplicity. We observed a significant increase in both SI (p=0.03) and colonic tumors (p=0.006) in the B6-Min/JGMHSD group as compared to the B6-Min/JGMJAX group. Additionally, we observed trending increases in SI and colonic tumor numbers in B6-Min/DGMHSD compared with B6-Min/DGMJAX mice. These data suggest that the GM has a critical role in the SI and colonic tumor phenotype. We also noted that B6-Min/D animals had more SI tumors than B6-Min/J animals within each GM group. To ensure that no known C57BL/6J modifiers existed in the population, founder animals were genotyped and found negative for the B6-Atp5a1Mom2 resistance allele, indicating that the Mom2 mutation is not responsible for the phenotypic differences between the two colonies. Importantly, there was no difference in colonic tumor numbers between groups with the same GM. Together, these data suggest that host genetic differences play a role in the SI phenotype, but not the colonic phenotype. We conclude that both the GM and host genetic factors within the C57BL/6J-ApcMin population contribute to the SI tumor phenotype. However, the GM primarily defines the colonic tumor phenotype. In addition to revealing highly important information regarding tumorigenesis in the ApcMin mouse, these results will have a critical impact on reproducibility of studies using this model.