Optimization of Induction and Culture Strategies for Calli with Stem Cell-like Features for the Production of Functional Natural Products from Korean Native Plants: Case Studies Involving Artemisia, Ixeridium, and Perilla

Plant stem cells and calli with stem cell-like features provide scalable, sustainable platforms with superior genetic stability, reduced aggregation, and improved yield consistency compared to dedifferentiated cells (DDCs).We implemented a four-step workflow-induction, biochemical and ultrastructural validation, suspension-culture optimization, and LC-MS/MS quantificationspanning three Korean native species: in A. capillaris, root cells generated from calli with stem cell-like features exhibited approximately fourfold higher artemisinin content than intact plants; in I. dentatum, calli with stem cell-like features demonstrated approximately tenfold greater chrysoeriol-7-O-glucoside accumulation and secreted more than twofold more of this metabolite into the medium than was present intracellularly; and in P. frutescens, calli with stem cell-like features yielded approximately threefold more rosmarinic acid compared to DDCs (40.4 1.4 vs. 12.9 0.7 mgg -1 ).At present, the integration of single-cell and spatial omics, elicitation, and advancements in media and scale-up engineering with AI technologies (ML soft sensors, digital twins, hybrid models) supports real-time process monitoring and control, minimizing variability and facilitating lab-to-industry translation.Future priorities include the development of robust stemness markers, standardized sensing strategies enabled by cross-species-generalizable AI, and long-term bioreactor reliability to achieve consistent, large-scale, and eco-friendly production of natural products using systems based on CMC-and calli with stem cell-like features.