Technical Abstract: Exposure to maternal obesity (MO) increases the risk of obesity in adult-life. MO was induced in rats by overfeeding via total enteral nutrition. Male offspring from obese rats gain greater body weight, fat mass and develop insulin resistance when fed high fat diets. However the mechanisms underlying MO-induced maternal programming of offspring metabolism remain unclear. Epigenetic changes such as altered methylation of CpG sites in the offspring DNA, following exposure to MO in utero, may be one mechanism leading to pleiotrophic effects. Genomic regions in which the content of CpG is high are referred to as CpG islands (CpGIs) and methylation of CpGIs is normally associated with transcriptional gene repression. In the present study, we analyzed hepatic DNA methylation in offspring of lean and obese rats. We employed a procedure to enrich methylated DNA using capture with methylbinding protein followed by Illumina sequecning (36-bp, single-read). Two pools of liver samples (3-4 separate offspring in each pool) were utilized for each group. Sequencing was carried out on a GAIIe sequencer and resulted in approximately 18 million reads per sample. Model-based analysis of ChiP-Seq data (MACS) was used to identify peaks using lean group samples as control and more than 4,000 peaks were identified. A program was developed to classify the regions under each of the peaks depending on their CpG content. Promoters in neighboring regions of high CpG content were identified. The results suggest MO may result in epigenetic differences in the offspring’s DNA resulting in metabolic alterations affecting energy utilization.