The global fermentation industry faces persistent bottlenecks in scaling laboratory innovations to industrial production, and the integration of synthetic biology (SynBio) and artificial intelligence (AI) within the Design–Build–Test–Learn (DBTL) loop has yielded inconsistent industrial outcomes. This review proposes that transformative impact requires a “scale-up-gated DBTL” framework, in which explicit decision gates constrain every iteration. At the Design phase, scale-down simulation data must inform genetic design choices. At the Test phase, downstream processing compatibility and industrial robustness metrics are enforced as non-negotiable evaluation criteria. At the Learn phase, techno-economic analysis (TEA) and life-cycle assessment (LCA) serve as the convergence criteria, replacing traditional titer plateaus. Through a qualitative cross-sectoral analysis of food, pharmaceutical, agricultural, and energy fermentation, the analysis reveals that workflows incorporating such constraints consistently bridge the valley of death, whereas unconstrained DBTL systematically converges on laboratory optima that are industrially unviable. Five strategic priorities are outlined—embedding TEA/LCA into DBTL, adopting scale-down simulation, building open fermentation data repositories, harmonizing regulatory frameworks, and fostering cross-disciplinary training—as prerequisites for progressing toward fully autonomous, scale-up-aware biomanufacturing.