Team:Goettingen/Measurement
Team Göttingen
iGEM 2018
Glyphosate on my plate?
| E-mail: | igem2018@gwdg.de |
|---|---|
| Adress: | c/o iGEM 2018 |
| Department of General Microbiology | |
| Grisebachstraße 8 | |
| 37077 Göttingen | |
| Germany |
We are especially grateful to our numerous sponsors on gofundme, who supported our project and made it possible.
Every public donor, who helped us to achieve our goals got a special place on our bacterial wall of fame and a handdrawn microbe to show our appreciation. A link to that wall can be found here:
Measurement
Glyphosate is a widely used herbicide with unknown risks for the environment and human health. Our goal is to develop a simple and cheap detection system, which allows average consumers to assess whether glyphosate is present in their food products. The colour-based glyphosate detection system relies on monitoring intraspecies competition of strains of the environmental bacterium Bacillus subtilis tolerating different amounts of the weed killer. To obtain B. subtilis strains that could grow with glyphosate, we have evolved the bacteria in the presence of the weed killer. These experiments revealed that B. subtilis rapidly adapts to glyphosate at the genome level by the acquisition of beneficial mutations, rendering the bacteria insensitive to the herbicide. The characterization of the mutants led to the discovery of the first glyphosate uptake systems! Furthermore, the bacterial strains tolerating different amounts of glyphosate facilitated the construction of our herbicide detection system.
Our measurement method uses fluorescence that is linked to competing Bacillus subtilis strains. The sensitive strain is tagged with GFP, while the glyphosate-resistant strain is tagged with mOrange. In a competition assay are both strains 1:1 mixed and grown with a possibly glyphosate-containing sample. The ratio between green and orange fluorescence gives information on the glyphosate concentration. The functionality is shown here.
Furthermore, we constructed another system that uses the activity of the β–galactosidase that is expressed in the wildtype-like strain BP193 (trpC2 palf4::lacZ). The glyphosate-resistant strain BP235 (trpC2 amyE::(morange-lacZ-aphA3) ΔgltT::spc ΔgltP::cat eliminates strain BP193 stronger the more glyphosate is present. This elimination can be monitored via the intensity of the blue color of the medium (Results). The intensity is compared to a test strip and the concentration can be read.
The functionality of our product is shown in the following picture (Figure 1).
Figure 1. Explaning the principle of our product.
Roundup® is sprayed on soil, Then, water is applied to solve the glyphosate from the sample. The solution is filtered through a filter devide to avoid any other contaminations like other microorganisms. The sample drops onto solid media powder with all necessary ingredients and Bacillus subtilis spores from the two competing strains. The sample is incubated for 10 h at 37°C with agitation. Now, the blue color can be compared to a test strip. The intensity of the blue color determines the concentration of glyphosate in the sample. The usage of our product is also shown in the following movie.