Inducible two-step cultivation of microalgae: engineering the growth and lipid biosynthesis for biofuel production

Microalgae have significant economic importance due to their fast growth and diverse biomass composition. However, achieving cost-effective large-scale production while maintaining the high yields of the desired metabolite remains a major challenge. Conventional cultivation strategies face an inherent trade-off: conditions that promote microalgae development often suppress metabolite synthesis, whereas stress conditions that induce the desired metabolite synthesis inhibit growth and overall biomass production. In recent years, there has been increasing interest in a two-step cultivation, in which microalgae are first grown under optimal conditions for maximal biomass, followed by exposure to stress to trigger targeted metabolite synthesis. Despite its advantages, this approach also faces limitations in precisely controlling metabolite induction and in avoiding undesirable stress responses. To overcome this issue, integrating inducible genetic switches in the second cultivation phase, particularly stress-responsive promoters linked to the gene of interest, offers a promising strategy to enhance metabolite production in a controlled manner. However, until now, there are no published studies on the utilization of inducible promoters along with a two-stage cultivation system for microalgae. The inducible systems are activated by specific stress factors or substrates, enabling targeted activation of desired pathways while limiting off-target effects. This review provides an overview of the mechanistic framework of the two-step cultivation system, types and mechanisms of inducible promoters, and switch-based metabolic regulation in microalgae. Finally, we will highlight the challenges and opportunities for integrating synthetic biology tools with cultivation engineering to enhance and sustain targeted metabolite production in microalgal biorefineries.