P.L.A.N.T. Plant-based Ligand Activated Noxious agent Tracker - make the invisible visible

Contaminated Sites

In Germany, we have a problem that has been left behind by past generations. Spread all over the country, there are known as well as unknown storage sites of chemical weapons from both world wars. The reason for this is the insufficiently documented and uncontrolled disposal of Chemical Weapons. Contaminated sites are suspected in over 200 locations in Germany. They pose a toxic threat to humans and the environment, if leakages occur.

Hydroculture

We decided to detect several chemicals using our system, namely methylphosphonic acid (MPA), diisopropyl methylphosphonate (DIMP), diethyl methylphosphonate (DEMP), thiodiglycol (TDG) and benzenetricarboxylic acid (BTCA). These chemicals, related to chemical weapons but safe to work with, are stable in nature and have the potential to severely contaminate the environment and groundwater. For our plant-based detection system of these chemicals, it is important to test if they are absorbed by plants and therefore are detectable. We cultivated Nicotiana benthamiana as hydroculture and demonstrated that the plants tolerate DIMP, DEMP, MPA and TDG in different concentrations. Furthermore, we showed the up take of said chemicals into the plant.

Protein Engineering

Crucial for the functionality of our detection system for degradation products of chemical weapons is a receptor which is able to bind the targeted chemical. By applying computational protein design on a ribose binding protein, we created a functional and specific receptor to bind a weapon related chemical. By using a modular system, the receptor can be changed easily to create a customized plant for detection of specific chemicals. One way to do this is the computational design we performed (Parts: BBa_K3900001, BBa_K3900002, BBa_K3900003, BBa_K3900004, BBa_K3900005, BBa_K3900006).

Reporter Ruby & Anthos

After detection of a chemical weapon degradation product and the activation of the signaling cascade, a visible output is created. Therefore, we introduce the RUBY reporter (BBa_K3900028), which comprises genes for betalain biosynthesis. Expression of this reporter results in a visible red coloration of the plant.

Bacterial and plant test systems

Our plant-system allows the detection of specific chemicals based on a signaling cascade activated by a receptor that specifically binds ligands. To perform initial tests with our signaling cascade and our newly engineered receptors, another fast growing and easy manipulative model organism was required. Therefore, we worked with Escherichia coli for first testing of a bacteria adapted signaling cascade and the engineered receptors. To demonstrate its functionality, we tested the plant signaling cascade by transient expression in Nicotiana benthamiana (Parts: BBa_K206000, BBa_K216004, BBa_B0014, BBa_K216003, BBa_K808000, BBa_R0082, BBa_E0040, BBa_P1003, BBa_K731722).

Human Practices

In order to develop and improve P.L.A.N.T., our plant based detection system for chemical degradation products, we were in constant contact with different experts and stakeholders throughout the year. With their help, we were able to adjust the aim of our application to the detection of degradation products for identification of contaminated sites and to further fine-tune our experiment project realization. When considering releasing a GMO, several views should be taken into consideration as well as the laws and regulations.