During our project, we had three smaller teams within our main team. Each of these teams had different but interlinked objectives. Below we have outlined the main achievements of those projects:
We successful generated a range of phytobricks that have been submitted to the registry and can be utilised by future iGEM teams who are interested in using the plant expression systems in their projects. Details of these phytobricks can be found on our Basic and Composite parts pages.
Team Luciferase created the 35S:Luc+:NosT construct , and quantified the activity of the 35S promoter using a luciferase reporter assay. These assays are detailed on the Part Experience page.
This image shows leaf discs taken from plants infiltrated with Agrobacterium containing the 35S:Luc+:NosT construct. The red regions show high levels of luciferase expression. This part can now be used by other teams in the future as a control when performing luciferase reporter assays.
We successful generated a range of level 1 phytobricks that included a plant promotor hooked up to TSHH. We attempted to use these constructs to express TSHH in tobacco leaves in a collaboration between Team TSH and Team Plant_P.
Constructs were introduced into tobacco leaves and after 2 days either salicyclic acid (GST/PR2) or jasmonic acid (PDF1.2) was also inflitrated into the leaves in order to induce gene expression. We performed crude protein extracts from these leaves and then compared to agrofiltrated leaves that had not been treated with the appropriate plant hormone. We used nickel beads to isolate the His-tag TSH protein and compared those samples to crude extracts in an attempt to identify bands that are specfic for the TSHH.
This protein gel showing the results of our protein extracts from leaves infiltrated with either GST:TSHH or PR:TSHH constructs. Following construct infiltration, they were then infiltrated with salicylic acid. A protein extract was carried out on the leaves to assess whether or not our TSHH antagonist was present. By comparing with the negative control, we could identify any specific bands corresponding to the TSHH protein. In further experiments we will attempt to optimise this expression and purification.
This protein gel shows the results of infiltrating PDF1.2:TSHH constructs, followed by infiltration of jasmonic acid. The lack of an obvious band in the 'beads' samples, and the fact there is no band unique to infiltrated leaves when compared to the negative control suggests that this experiment did not work as hoped.
We generated a model that investigated the potential use of the tobacco leave expression system for the purification of human therapeutic agents. Our model is outlined on our modeling page.
We conducted a survey for almost 300 participants that assessed public opinion on the use of genetic modification for the production of pharmaceuticals in animals or plants. We also discussed with a range of experts the scientific and economic advantages of using plants for protein production. This allowed us to perform an analysis on the benefits of the plant expression system, which integrated with our modelling work. This can be found on Our Research page.
We are very proud of maintaining our lab notebook every day of the 10 weeks that we were in the lab. This can be found on our Diary page.