Objective To explore the most suitable transplant site for islet transplantation by comparing the efficacy of gastric subserosal transplantation and portal vein islet transplantation in the treatment of diabetic rats. Methods Streptozotocin-induced diabetic rats were divided into gastric subserous group (n = 6), portal vein group (n = 6) and control group (n = 6). Type XI collagenase was injected through choledoch to digest the pancreas, and the islets were isolated and purified. The specificity of the islet was judged by dithizone (DTZ) staining, and IEQ and islets purity were counted. The activity of islets was determined by trypan blue test. 500IEQ fresh islets were transplanted into the gastric subserosa and into the portal vein, respectively. The control group was infused with the same amount of RPMI 1640. After islet grafting in the diabetic rats, the changes in blood glucose and intraperitoneal glucose tolerance were evaluated to assess the function of islets. Results The number of islets was (310 ± 42) in each transplanted rat, and the purity of islets was (89 ± 4.52)% obtained by DTZ staining and the activity of islets was (89.08 ± 3.76)% obtained by trypan blue staining. The glucose levels in the gastric subserous and portal vein groups were significantly decreased after transplantation compared with those before transplantation (P < 0.05) and significantly different from that in the control group (P < 0.05). After transplantation the average blood glucose level of the gastric subserous group was significantly lower than that of the portal vein group (P < 0.05). The time of effective control of blood glucose in the gastric subserous group and the portal vein group was (18.0 ± 1.9) d and (11.6 ± 2.5) d (P < 0.05). The rats of the gastric subserous and portal vein groups had good intraperitoneal glucose tolerance. Conclusions In the short term, the effect of gastric subserous islet transplantation for treatment of rat diabetes is better than that of portal vein islet transplantation. Compared with portal vein islet transplantation, the operation of gastric subserous pancreatic islet transplantation is simple and safe, and gastric subserosa can be considered as one of the most suitable parts of islet transplantation.