<p>Adventitious root cultures have attracted significant interest as a stable and sustainable in vitro system for the enhanced production of pharmaceutically valuable secondary metabolites in medicinal plants. These roots, which are developed from a non-root tissue under exogenous plant growth regulators’ control, have numerous advantages over classic growth and culture systems, such as high rates of growth, stability of their genes and biochemistry, and ability to efficiently function in high-biomass scale bioreactor systems. Adventitious root cultures’ induction and multiplication are determined by several variables, which include the level and concentration of auxins, composition of the medium for growth, and quality of the explant used. Additionally, the use of elicitors and environmental factors have also been identified as highly important means to increasing the yield of metabolites such that adventitious root cultures remain a valuable choice towards commercial exploitation. Recent developments in molecular biology have also helped to elucidate the fundamental signaling pathways and patterns of gene expression involved in adventitious root development and secondary metabolism. This review presents a systematic overview of the physiological, biochemical, and technological facts of adventitious root culture systems and their use in secondary metabolite biosynthesis. The possibilities of industrial-scale use and compatibility with metabolic engineering practices are also discussed, highlighting the importance of adventitious root cultures in contemporary plant biotechnology and pharmaceutical production.</p>