Generating Strains

I have spent most of the past month beginning to engineer my Trojan Horse strain. After many rounds of trial and error, I have successfully used PCR to amplify a toxin cassette. I am now in the process of transforming several strains to express the toxin. Once this is done, I will assay toxin production against a known sensitive strain. This has been a slow process so far, but I now have at least one potentially successful transformation. Once I confirm toxin production, I will be able to begin experiments competing my toxin-producing strains against other strains and testing whether they can infiltrate and disrupt biofilms.

I am also exploring how to inject an inducing agent into an established colony, as I ultimately aim to generate my Trojan Horse strain to express a toxin under an inducible promoter. To visualize the effectiveness of different administration techniques, I am engineering strains to express fluorescence under the inducible galactose promoter. Injecting colonies of these strains with galactose should induce fluorescence by cells that the galactose reaches, showing me how well the inducing agent is penetrating the colony. I will use this visual aid to develop an effective administration method that I can use with my Trojan Horse strain.