Metabolic engineering and synthetic biology employing Lactococcus lactis and Bacillus subtilis cell factories

[Display omitted] •Modifying metabolic fluxes in L. lactis and B. subtilis to obtain good production hosts for food, pharma, and biotechnology.•Overview of synthetic biology tools for L. lactis and B. subtilis for the construction of custom-made microbial chassis.•Eukaryotic protein expression in L. lactis for biomedical applications. Metabolic engineering and synthetic biology approaches have prospered the field of biotechnology, in which the main focus has been on Escherichia coli and Saccharomyces cerevisiae as microbial workhorses. In more recent years, improving the Gram-positive bacteria Lactococcus lactis and Bacillus subtilis as production hosts has gained increasing attention. This review will demonstrate the different levels at which these bacteria can be engineered and their various application possibilities. For instance, engineered L. lactis strains show great promise for biomedical applications. Moreover, we provide an overview of recent synthetic biology tools that facilitate the use of these two microorganisms even more.