Our Sensor

The goal of our project is to design a biosensor to rapidly identify cattle known as super shedders. Super shedders are cattle that excrete extremely large numbers of E.coli O157:H7 a subgroup of E. coli referred to as Enterohemorrhagic E. coli (EHEC). EHEC organisms produce a toxin called Shiga toxin or verotoxin. This toxin binds to blood cells and lyses them resulting in hemolytic-uremic syndrome (HUS) in humans and can be deadly. Supershedders excrete in the range of 10 ⁷ to 10 ¹⁰ E. coli O157:H7 exceeding normal cattle by 3-6 logs and can contaminate other cattle in the same holding pen as well as the meat downstream. Super-shedders are thought to be the reason for 95% of the E. coli O157:H7 contamination in the industry.

We are building a DNA-based biosensor that specifically detects the gene called stx2. Our device will be useful for the detection of not only E.coli O157:H7 but also other EHEC strains. Click on the components below to learn more about their design and function. View our animation below to see how the system would actually work!

The FerriTALE System:

As seen in the video above our goal is to develop a strip based assay that can alert us upon detection of pathogenic DNA sequence, namely Stx 2. We will treat our sample collected from the supershedders with a TALE-Ferritin complex (a FerriTALE!). Then this DNA-TALE-Ferritin complex would be flown over our strip which contains the second TALE that recognizes yet another 18-20 bp region on the stx 2 gene. This second TALE will immobilize our DNA-TALE-ferritin complex on the strip. Following immobilization we will add substrate to our strip. Only in the presence of both the TALEs we will see a colour change on the strip.

A Platform Technology for iGEM

TALEs are modular proteins for which the amino acid to nucleotide binding code has been solved and the TALEs therefore have a great potential to be used as a platform technology. Since the amino acid sequences which bind to the nucleotides are known we can engineer the DNA binding domains to detect practically any DNA of interest with high specificity.

We are using TALE proteins as our sensory element. TALEs are very powerful tools since they can be engineered to bind to any 12 to 31bp sequence of DNA (source: Golden Gate TALEN and TAL Effector Kit 2.0). Therefore, by just swapping the TALEs in our PLASMID XYZ, one can detect any DNA of interest. To further ease this process, we have incorporated a KasI restriction cut site at the end of all TALEs in all the constructs. Using the KasI restriction enzyme and one of the biobrick prefix enzymes one can incorporate their own engineered TALE into the part, and use it for detecting their DNA of interest.