Diet-induced obesity (DIO) has been shown to increase DNA methyltransferase (DNMT) expression and DNMT binding at obesity-associated genes. Natural compounds have the potential to reverse obesity-associated gene expression via the regulation of DNA methylation. The objective of this study was to determine the effect of the bioactive compounds of flaxseed on DNMT and obesity-associated gene expression. Sixty C57BL/6J male mice were randomly assigned into one of the following diet groups and fed for eight weeks: 45% kcal fat; 45% kcal fat, 10% whole flaxseed; 45% kcal fat, 6% defatted flaxseed; 45% kcal fat, 4% flaxseed oil; and 16% kcal fat. DNMT1, DNMT3a, DNMT3b, leptin, and peroxisome proliferator-activated receptor (PPAR)-α expression levels in muscle and adipose tissues were determined by real-time PCR. The results indicate that bioactive compounds of flaxseed affect selected gene expression. Further research is needed to identify the specific mechanisms regulating leptin or PPAR-α expression during DIO development.