TraDIS as a tool for accelerated lab-based evolution

The use of long term laboratory experiments coupled with whole genome sequencing is currently proving a valuable tool for understanding the relationship between genotype and phenotype. Previous studies in our lab have shown that in replicated long term evolution at low pH, using E. coli as a model organism, mutations arise repeatedly in the same key regulator genes, and we have shown using strain construction and competition experiments that these are responsible for contributing the most to fitness of the evolved strains. Often, though not always, we were able to show that these were loss of function mutations. This project will study evolution of stress resistance under laboratory conditions in more detail by investigating further the contributions of individual genes to the fitness of the evolved strains and determining the mechanisms by which they act. One major issue with long term evolution is the length of time taken to conduct the experiment. We will therefore look at the potential of using high throughput methods for speeding up the generation and analysis of data from lab-based evolution experiments. If this approach is successful we will look at the roles of different parameters to see how robust the results are to parameter variation including the pH, the specific acid used, the media used, the strain genotype, the dilution rate, etc. This will enable us to link our data to more realistic "real world" scenarios and will be useful for informing future studies using lab-based evolution, for example for more efficient strain engineering or for studying emergence of resistance traits that might be important in organisms that cause food spoilage or infection.