Ileal neobladder construction is a common treatment for patients with bladder cancer after radical cystectomy. However, metabolic disorders caused by transposed bowel segments occur frequently. Bladder tissue engineering is a promising alternative approach. Although numerous studies have reported bladder reconstruction using acellular and cellular scaffolds, there are also disadvantages associated with these methods, such as immunogenicity of synthetic grafts and incompatible mechanical properties of the biomaterials. Here, we engineered an autologous peritoneal graft consisting of a peritoneal sheet and the seromuscular layer from the ileum. Three months after the surgery, compared with the neobladder made from the ileum, the reconstructed neobladder using our new method showed normal function and better gross morphological characteristics. Moreover, histopathological and transcriptomic analysis revealed urothelium-like cells expressing urothelial biomarkers appeared in the neobladder, while no such changes were observed in the control group. Overall, our study provides a new strategy for bladder tissue engineering and informs a variety of future research prospects.