Since DNA methylation plays a causal role in pancreatic remodeling and development, thus modulation of this epigenetic mechanism is essential in treatment of diabetes. This provoked us to examine the effect of a known anti-diabetic agent, an isolated polysaccharide, on the methylation pattern of Ins-1 and Pax-4 in diabetic rats. Here, a polysaccharide fraction was isolated from Rosa canina and analyzed using NMR, FTIR and MS/MS techniques. Diabetes was established by using intraperitoneal injection of STZ in male Wistar rats. After treatment, pancreas was removed and DNA was extracted and bisulfite treated by a DNA methylation kit. PCR and real-time PCR were used to determine the levels of methylated and/or unmethylated Ins-1 and Pax-4 genes. The levels of blood glucose and weight body were normalized in diabetic rats exposed to isolated polysaccharide. The level of unmethylated Ins-1 was upregulated in diabetic rats which is downregulated in metformin and polysaccharide-treated ones. In diabetic rats, the content of methylated Pax-4 was increased while it was decreased in polysaccharide-treated group. Interestingly, the methylation pattern of Pax-4 in metformin group was the same as diabetic ones. Data clearly indicated that polysaccharide can reduce the level of blood glucose by modulating the methylation pattern of Pax-4 and Ins-1.  This study sheds light on the importance of DNA methylation modulation as a promising therapeutic strategy in diabetes.