UC San Diego

Engineering novel, biological functions with synthetic genetic circuits is easier than ever before due to constant discovery of new genetic parts, development of efficient ways to put those parts together, and innovative methods to model and study complex phenotypes. As the field of Synthetic Biology continues to grow, different focus areas and challenges have come to light. In this dissertation, I identify key themes and current challenges in Synthetic Biology and use novel circuit design and screening approaches to build on existing research in these areas. One growing area of focus is controlling and coordinating microbial behavior at the population level. A major challenge in this area is screening dynamic, population-level circuits for desired behavior with the same throughput as single-cell circuit screening methods. Chapter 2 of this dissertation introduces a novel microfluidic device that enables screening of mutant bacterial libraries for complex, population-level phenotypes that change with time. Chapter 3 focuses on genetic circuit screening as well, but from a more application-based standpoint. Specifically, a workflow is developed for screening bacterial expressed toxins for their ability to inhibit cancer cell growth when released via engineered bacterial cell lysis. Novel candidates are uncovered for bacterial-based cell therapies. While noise and heterogeneity are typically avoided in synthetic biology, Chapter 4 presents a novel strategy for exploiting engineered heterogeneity in bacterial populations to enable the populations to quickly adapt to varying environments. Chapter 5 addresses the need for new, quantitatively-characterized inducible promoter systems in yeast. The systems developed in this chapter are designed to be compatible with existing modular cloning toolkits in yeast, making their implementation simple and standardized. Together, the results presented in this dissertation demonstrate novel strategies for controlling population-level behavior in bacteria, screening genetic circuits for complex phenotypes, and creating new genetic parts for circuit regulation.