Folate is an essential water soluble B-vitamin required for the de novo synthesis of purines and dTMP and the synthesis of methionine. Folate intake,, generally folate deficiency, has been associated with CRC and ALL risk. Here, we used the MutaMouse model to determine the mutagenic potential of dietary FA in the colon, and to determine if there is a tissue- and diet-specific effect induced by FA intake in colon and bone marrow. Male mice were fed experimental FA defined diets (deficient, control, supplemented) for 20 weeks from weaning. NGS was used to sequence the lacZ reporter gene and we determined the FA-induce mutation spectra in both tissues. Here, we demonstrated, for the first time, the mutagenic potential of FA-intake, the FA-induced mutation spectra by different FA intake levels and that the mutagenic potential of FA is tissue-specific.