(646a) Development of Synthetic-Hybrid Systems for Enhanced Gene Expression Control

Authors: 
Forkus, B., University of Minnesota
Kaznessis, Y. N., University of Minnesota

Development of synthetic-hybrid systems for enhanced gene expression control

Brittany Forkus and Yiannis Kaznessis

Controllable gene regulatory systems that exhibit new biological phenotypes can be developed by arranging naturally occurring regulatory genes in synthetic DNA sequences. In our work, we have built several synthetic DNA promoter systems by stitching together various components of the common and well-studied tetracycline, luminescence, and lactose operons.

Previously we designed simple logical AND gates by shuffling tetO and lacO operator sites in a single transcriptional unit. We then built ProteOn, the first inducible activator transcription factor, by fusing the tetracycline repressor with the activating domain of the luxR protein.  

In this presentation we will discuss the most recent system we built, which implements inducible activation of protein expression with positive-feedback. We call this new system ProteOn+. We characterized this system extensively by expressing green fluorescent protein (GFP), and using flow cytometry techniques. We compared ProteOn+ to commercially available promoter systems and found that ProteOn+ elicits many-fold increases in gene expression than commonly used DNA promoters. We believe that ProteOn+ adds significant value to the current synthetic biological- toolbox for bacterial gene expression.

In this presentation we will also discuss applying ProteOn+ in the development of antibiotic technologies. We have engineered probiotic E. coli strains to express antimicrobial peptides from ProteOn+. We assess the strength of this synthetic DNA promoter and its utility in the delivery of antimicrobials to treat gastrointestinal infections.