Master of Science, Miami University, 2024, Chemical, Paper and Biomedical Engineering

Engineering microbes for sustainable bioproduction is a promising avenue to tackle problems in pollution, medicine, and energy. Traditional model organisms have received much attention due to their easily manipulated genetics and well characterized growth. However, there are a wealth of non-model microbes with attributes that are beneficial for biotechnological applications, but genetic tools must first be developed to engineer them by incorporating foreign DNA and characterizing bioparts for controlled expression. In this work, we are looking to expand the genetic toolbox for an environmental-isolate of Priestia megaterium, called SR7, that can survive under high pressures of CO2. Bioprocessing under high pressures of CO2 is expected to simultaneously create an aseptic culture environment as well as extract bioproducts as they are formed, both benefits for continuous bioproduction. An enhanced transformation method was developed, permitting bioparts characterization. We have compared promoters, origins of replication, and antibiotic resistance genes between two plasmid architectures. We identified three functional Gram-positive origins of replication for SR7. Three plasmids were verified to have enhanced recombinant protein production compared to a previously used plasmid.

Committee: Jason Boock (Advisor); D.J. Ferguson (Committee Member); Jason Berberich (Committee Member) Subjects: Biology; Chemical Engineering; Microbiology; Molecular Biology