Exploration of Epigenetic Regulation and Development of Global and Targeted Epigenetic Engineering Tools for Taxus Plant Cell Culture

Microbes play critical roles in mediating the health of soil and plants. Synthetic biologists have developed a variety of production, sensing, and response capabilities in living microbes; however, these tools currently cannot be implemented in the environment due to the risk of uncontrolled proliferation of engineered microbes. To address this issue, my lab has been working on designing strategies for intrinsic biological containment of microbes, with a focus on the technique of synthetic auxotrophy. A synthetic auxotroph is an organism engineered to depend on the provision of synthetic nutrients not found in nature. This poster highlights our latest characterization of Escherichia coli synthetic auxotrophs, which includes continuous evolution and direct addition to mammalian cell cultures. It also describes our successful efforts to engineer the incorporation of synthetic amino acids in the gram-positive microbe Bacillus subtilis. B. subtilis is a model system for both the study of bacterial cell biology and for industrial uses as a probiotic, rhizobacterium, efficient protein secretor, and spore-former. To enable greater understanding and control of proteins in B. subtilis, we achieved incorporated many distinct synthetic amino acids within proteins. We used these systems to achieve click-labelling, photo-crosslinking, and translational titration. Synthetic auxotrophy in E. coli and eventually in B. subtilis could serve as an enabling safeguard for engineered plant probiotics, particularly those with pre-programmed limited lifetimes in the environment.