Antibiotics and their corresponding resistance genes act as a tool to control bacterial survival. Antibiotic resistance is used to select for desired engineered cells, and study how pathogens acquire resistance to continue infection. Here, we develop tools to control the expression of antibiotic resistance genes using light. To accomplish this, we use optogenetics, the regulation of cellular behavior using light as a direct and programmable input for gene expression. We develop an optogenetic recombinase in Escherichia coli through split-protein engineering techniques, and characterize the behavior of our best candidate in detail: a split Cre recombinase that responds to blue light. We apply this optogenetic system to control the expression of resistance genes for four antibiotics: ampicillin/carbenicillin, kanamycin, chloramphenicol, and tetracycline. By varying the expression levels of these genes, we tune the concentrations at which bacteria can survive before and after light exposure. We then apply this system to improve production of fatty acids. Finally, we make progress toward characterizing the impact of resistance activation timing on bacterial survival. This work creates tools that are broadly useful for spatiotemporal control of bacterial survival, and enables precise studies on how bacterial resistance spreads at the single-cell level. / 2024-08-29T00:00:00Z
| Identifer | oai:union.ndltd.org:bu.edu/oai:open.bu.edu:2144/46644 |
| Date | 30 August 2023 |
| Creators | Sheets, Michael Brian |
| Contributors | Dunlop, Mary J. |
| Source Sets | Boston University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
| Rights | Attribution-NonCommercial-ShareAlike 4.0 International, http://creativecommons.org/licenses/by-nc-sa/4.0/ |
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