Sweetpotato (Ipomoea batatas (L.) Lam.) holds a crucial position as one of the staple foods globally, however, its yields are frequently impacted by environmental stresses. In the realm of plant evolution and the response to abiotic stress, the RNA helicase family assumes a significant role. Despite this importance, a comprehensive understanding of the RNA helicase gene family in sweetpotato has been lacking. Therefore, we conducted a comprehensive genome-wide analysis of the sweetpotato RNA helicase family, encompassing aspects such as chromosome distribution, promoter elements, and motif compositions. This study aims to shed light on the intricate mechanisms underlying the stress responses and evolutionary adaptations in sweetpotato, thereby facilitating the development of strategies for enhancing its resilience and productivity. 300 RNA helicase genes were identified in sweetpotato and categorized into three subfamilies, namely IbDEAD, IbDEAH and IbDExDH. The collinearity relationship between the sweetpotato RNA helicase gene and 8 related homologous genes from other species was explored, providing a reliable foundation for further study of the sweetpotato RNA helicase gene family's evolution. Furthermore, through RNA-Seq analysis and qRT-PCR verification, it was observed that the expression of eight RNA helicase genes exhibited significant responsiveness to four abiotic stresses (cold, drought, heat, and salt) across various tissues of ten different sweetpotato varieties. Sweetpotato transgenic lines overexpressing the RNA helicase gene IbDExDH96 were generated using A.rhizogenes-mediated technology. This approach allowed for the preliminary investigation of the role of sweetpotato RNA helicase genes in the response to cold stress. Notably, the promoters of RNA helicase genes contained numerous cis-acting elements associated with temperature, hormone, and light response, highlighting their crucial role in sweetpotato abiotic stress response.