Based on the data of A-share listed manufacturing companies and urban panel data from 2006 to 2023, this paper empirically examines the impact and mechanisms of manufacturing transformation and upgrading on regional scientific and technological innovation capacity. The research finds that the transformation and upgrading of the manufacturing industry promotes the regional scientific and technological innovation capacity, with both the agglomeration of innovation resources and the knowledge spillover effect serving as mediators. This presents a chain mediation structure from manufacturing transformation and upgrading to agglomeration of innovation resources, and then to regional innovation capacity through knowledge spillover effect. In addition, market demand and financial support exhibit non-linear moderating effects. Therefore, efforts should be made to continuously advance manufacturing transformation and upgrading, supported by increased investment in science and technology, well-regulated market demand and appropriate financial support, to enhance regional scientific and technological innovation capacity.