Teaching Synthetic Biology for Recombinant Protein Production: development of laboratory course for MSc students in Industrial Biotechnology

Recombinant protein production is a crucial expertise for biotechnology students, regardless of the production scale the student and future professional biotechnologist will face. For this reason, we have developed a course for MSc students in Industrial Biotechnology blending frontal lectures and laboratory activities. Frontal lectures aim to give the theoretical understanding of synthetic biology tools; while practical sessions engage the students to generate a GRAS yeast secreting a recombinant protein. Here, we present the vision, the structure, and the results of the course, wherein the students learn to use bioinformatics tools to design vectors and expression cassettes based on the Easy-MISE toolkit (Maestroni et. al., 2023) for gene integration in Saccharomyces cerevisiae exploiting the CRISPR-Cas9 system. Thanks to the modularity of the toolkit, students test the effect of different promoters and of different genome integration loci on the expression kinetics of a secreted recombinant laccase from the white-rot fungus Trametes polyzona (Bucchieri et al., 2024). The performance of the different S. cerevisiae strains generated is tested in E-flasks and is scaled-up to 2 L controlled bioreactors, where the production process is followed over time monitoring the enzyme activity. We believe that such a course is an essential experience for future industrial biotechnology professionals, providing an exhaustive overview of the entire design-build-test scheme and the basis for troubleshooting extracellular protein production processes in eukaryotic microorganisms.