Towards green biomanufacturing of high-value recombinant proteins using promising cell factory: Chlamydomonas reinhardtii chloroplast

Microalgae are cosmopolitan organisms in nature with short life cycles, playing a tremendous role in reducing the pressure of industrial carbon emissions. Besides, microalgae have the unique advantages of being photoautotrophic and harboring both prokaryotic and eukaryotic expression systems, becoming a popular host for recombinant proteins. Currently, numerous advanced molecular tools related to microalgal transgenesis have been explored and established, especially for the model species Chlamydomonas reinhardtii (C. reinhardtii hereafter). The development of genetic tools and the emergence of new strategies further increase the feasibility of developing C. reinhardtii chloroplasts as green factories, and the strong genetic operability of C. reinhardtii endows it with enormous potential as a synthetic biology platform. At present, C. reinhardtii chloroplasts could successfully produce plenty of recombinant proteins, including antigens, antibodies, antimicrobial peptides, protein hormones and enzymes. However, additional techniques and toolkits for chloroplasts need to be developed to achieve efficient and markerless editing of plastid genomes. Mining novel genetic elements and selectable markers will be more intensively studied in the future, and more factors affecting protein expression are urged to be explored. This review focuses on the latest technological progress of selectable markers for Chlamydomonas chloroplast genetic engineering and the factors that affect the efficiency of chloroplast protein expression. Furthermore, urgent challenges and prospects for future development are pointed out.