The mechanisms of weight loss and metabolic improvements following bariatric surgery in skeletal muscle are not well known, however epigenetic modifications are likely to contribute. The aim of our study was to investigate skeletal muscle DNA methylation after weight loss induced by Roux-en-Y gastric bypass (RYGB) surgery. Overall design: Muscle biopsies were obtained basally from seven insulin-resistant obese (BMI >40 kg/m2) female subjects (45.1±3.6 years) pre and 3 months post-surgery with euglycemic hyperinsulinemic clamps to assess insulin sensitivity. Four lean (BMI <25 kg/m2) females (38.5±5.8 years) served as controls. We performed reduced representation bisulfite sequencing next generation methylation on DNA isolated from the vastus lateralis muscle biopsies.