Demonstrate

Being a dual-aspect project, we had two sets of goals to achieve. Firstly, we aimed to produce an effective aphid insecticide using our siRNA constructs. Secondly, like previous Cardiff iGEM teams, we wanted to improve the PhytoBrick registry through the addition of significant parts, and through characterisation of widely used existing parts.

While we did not have time to test our siRNA constructs on aphids, we have demonstrated that they should work in theory, through our bioinformatic analysis. However, as we do not know how toxic they are, we cannot say that they will be an effective insecticide. Further work would allow us to establish this.

We did, however, succeed in our second criteria. We managed to expand the PhytoBrick registry, demonstrated by our two new reporter genes, GUS and mCherry, and a new plant promoter, RTBV, all of which can be seen on our results page. Using GUS as a reporter gene, we managed to successfully characterise the expression levels of the 35S CaMV promoter, and the NoS, G7, and 35S terminators, all of which are commonly used in plant synthetic biology. From these results, we would suggest that researchers should use the G7 terminator, as it has the highest and most consistent expression according to our experiments, though more replicates would be needed to completely confirm this. Our three best GUS assays highlight all of these achievements, shown below. Full documentation of these successes can be seen on the results page.

Figure 1: Results using the GUS reporter gene. Using the GUS reporter gene, we managed to characterise the 35S CaMV and RTBV promoters (left). This showed that both have constitutive expression, but that the 35S promoter produces stronger gene expression. We also characterised the influence of the nopaline synthase, G7, and 35S CaMV terminators on gene expression, all with the 35S CaMV promoter (centre and right). These results suggest that the G7 terminator contributes to the strongest and most consistent gene expression.