PROGRAMMING SELF-DIRECTED EVOLUTION IN BACTERIA

Abstract

Synthetic biology seeks to harness genetic design principles of living systems to re-engineer them for useful functions. Pioneering efforts in the field have inspired many exciting applications in bio-computation, metabolic engineering, and disease treatments. Novel technologies aiming to advance the throughput and timescale of synthetic biology projects hold the promise of driving innovations in biotechnology and biomedicine. The work in this thesis describes efforts to realize this vision. Specifically, a novel genetic platform was developed to implement self-directed evolution in living cells for facile generation and identification of beneficial biological mutants. Also, this platform facilitated adaptation of automated workflows in experimental evolution procedures to improve their success rate and scale-up potential. In this study, various genetic design strategies were devised and applied to create a reliable framework for future development of new genetic circuits aimed at similar purposes. In Chapter 2 and 3, critical genetic parts were identified and integrated into robust genetic devices to facilitate the construction of higher-order genetic circuits. In Chapter 4, these devices were assembled to create a proof-of-concept autoregulated genetic platform that can drive self-directed evolution of acid-tolerant phenotypes in Escherichia coli (E. coli). Chapter 5 discusses future research directions based on these results.