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Plasmid Design

Green light inducible plasmid

Red light inducible plasmid

Chromophore producing plasmid

Project Design

In-Vitro Selection

Genetic evolution by natural selection has guided life as we know it through billions of years of Earth’s harshest environments, giving rise to millions of diverse and steadfast lifeforms that we see around us today. In the lab, we can mimic this grandiose process to our advantage on a microscale using a technique called in-vitro selection¹. By following Darwin’s principles of natural selection in a controlled environment, it is possible to artificially evolve large groups of DNA molecules from randomness and select for those with useful functions. This is done by exposing large, “random libraries” of short DNA strands to other molecules of interest, and removing the species in the pool that don’t react to them in the way we desire. These reactions are observed on the macroscale using the tried and true technique of polyacrylamide gel electrophoresis on the radio-labelled, denatured (untied and linearized) DNA libraries after exposure. In the end, an assortment of DNA molecules comprising of more than a quintillion random nucleotide sequences is reduced to a handful of highly specific, functional product species.

If selected successfully to process a substrate on que, these nucleic acids are called DNAzymes. We work with DNAzymes that have been artificially selected to serve as a detection platform for E. coli, one of the most infamous antimicrobial drug-resistant strains of bacteria in the world^2,3. Our DNAzymes react in the presence of a cocktail of molecules radiated specifically from the extracellular matrix of the E. coli bacterium. The ensuing process results in the cleavage of an RNA unit bridging a quencher-fluorophore DNA complex, resulting in the emission of a signature green glow. In essence, we’ve used one of nature’s oldest solutions to generate something innovative, creative, and potentially revolutionary to the field of medical point-of-care testing.

[1] Wilson, D. S. & Szostak, J. W. In Vitro Selection of Functional Nucleic Acids. Annual Review of Biochemistry 68, 611–647 (1999).

[2] Aguirre, S., Ali, M., Salena, B. & Li, Y. A Sensitive DNA Enzyme-Based Fluorescent Assay for Bacterial Detection. Biomolecules 3, 563–577 (2013).

[3] Antimicrobial resistance: global report on surveillance. (World Health Organization, 2014).

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