Experiment Goals

The ultimate goal of this research is to create a novel biological system that ensures that the demineralization threshold is never crossed in the oral biome. In order to do this, one of the native bacteria, Veillonella parvula, would have to be engineered to: (1) detect and respond to pH drops due to lactic acid and (2) catabolize lactic acid in order to negate a drop in pH. Since V. parvula naturally contains lactic acid catabolic pathways, the detection and response aspect of this system will be the scope of this project. Our experiment has two intermediate goals: (1)Develop a new pH-responsive promoter and (2) construct a dual-state promoter with this new promoter.

The standardization inherent to the synthetic biology field means that genetic manipulation of a model system can in theory be easily translated to other species with the same genetic framework. Since the genome of Escherichia coli is well characterized, it will be used as the model system for our experiment.

This technology has the potential to dramatically change the oral health industry, but it also has applications to any biological system where pH monitoring would be necessary, including water sampling, food processing, and many other industries.

Fluorescence Assays

orescence assays of GFP expression will be conducted to analyze the activity of both the single promoters and the dual-state combinations. This will done by using a plate reader to measure the fluorescence per optical density of cells in minimal media with pH ranging from 7.5 – 3. The reason optical density will be taken into account is to control for fluorescence differences that may be based on different cell counts across trials. Instead the chosen measurement will be analogous to fluorescence per cell, giving much greater insight into the activity of the dual-state promoter in each cell at each pH level.