2017: GLOWzyme – An RNA-cleaving Fluorogenic DNAzyme Probe for Simple Detection of Pathogens

Current point-of-care diagnostics for bacterial infections are costly, time-consuming, and offer limited strain specificity. These challenges contribute to improper antibiotic usage, accelerating the propagation of antimicrobial-resistant (AMR) bacteria. We are developing DNAzymes – catalytically active ssDNA generated via in vitro selection – to serve as inexpensive and sensitive probes for the rapid detection of AMR bacteria. In the presence of targeted strains, the DNAzyme cleaves a fluorophore-RNA-quencher motif at the RNA site, generating fluorescence. As a proof-of-concept, we have adapted a known E. coli K12 DNAzyme for use in a plate-based assay, and are generating a novel DNAzyme to detect resistant strains of C. difficile. Simultaneously, we are leveraging machine learning techniques to predict potential DNAzymes, and are developing kinetic models to describe DNAzyme behaviour. Our project addresses the need for novel approaches within AMR detection and active antimicrobial stewardship – issues widely recognized by the experts in this field.
Learn more about the 2017 project here.

2016: Tumour-Seeking Bacteria

Gastrointestinal (GI) cancers have the highest cancer mortality rate in Canada due to difficulties in tumour diagnosis and treatment. Common anti-cancer therapies used today can lack specificity and have off-target effects, damaging the body in the process. Our focus is to engineer commensal Lactobacillus bacteria to bind and aggregate towards Her2+ GI tumours. Membrane-anchored binding proteins specific to Her2 will be engineered into Lactobacillus to redirect bacteria for aggregation at the tumour site. With sufficient bacterial density, a quorum sensing mechanism triggers production of a vital T-cell activating cytokine, interleukin-2 (IL-2), to recruit tumour-specific T cells for an anti-cancer response. Simulations of the bacteria in a possible tumour environment will help drive quorum-sensing design (e.g. promoter sensitivity). Ethical concerns were addressed through research into the feasibility of such a treatment, particularly into methods to minimize health risks, make the treatment financially accessible, and abide relevant legislation.
Learn more about the 2016 project here.

2015: Automating Protein Production Using Multichromatic Light

Light-based bacterial protein expression systems have been well documented. This is where you will find no deposit slots keep winnings. Follow the link! Tabor et al.demonstrated a multichromatic protein expression system in E. coli through the combination of the CcaR-CcaS green-light sensitive construct derived from cyanobacteria and a previously characterized red-light sensitive system. This could be leveraged for the expression of multiple genes by exposing the bacteria to different wavelengths of light, i.e. red and green. We propose the application of this construct to recombinant protein expression. Using genetic cloning and recombinant protein techniques, E. coli is concurrently transformed with three plasmids: a chromophore, a red-light sensitive system, and a green-light sensitive system. Protein expression is induced through shining red light on the bacterial population. Following this, cell lysis is triggered by shining green light, which is mediated by a T4 Holin/Endolysin system. This releases the protein of interest into the cell media, allowing for straightforward collection and purification. This system would have important applications in both research and industry, allowing for the optimization and potential automation of heterologous bacterial protein production.
Learn more about the 2016 project here.