Team:Imperial College/Composite Part
In this section we aim to present all the composite parts that we have built and used thorough our project. In addition we introduce our best composite part: the pBAD-Gp2 construct, which provides a novel non-lethal way of inhibiting the growth of bacteria.
This year our best composite part is the pBAD-Gp2 construct. It provides a new way to repress growth in bacteria without killing the host cell. We have demonstrated that the cell is able to recover its initial growth rate once Gp2 is no longer expressed and can recover back to a normal growth rate. We are excited to provide this new growth regulator for the synthetic biology community and for future iGEM teams.
In the presence of arabinose (the promoter was active from 100 μM L-arabinose), the araC transcriptional regulator activates the pBAD promoter, resulting in the production of Gp2, a protein from T7 phage. Gp2 binds reversibly to the bacterial RNA polymerase, preventing protein production, and preventing the cell from dividing.
Figure 1: Schematic of the pBAD-Gp2 construct
The part was created by cloning a synthesised sequence block that contained Gp2, an RBS and the biobrick prefix and suffix, into an pBAD construct created by the 2014 Imperial iGEM team.
The construct effectively downregulates the growth of E. coli in response to arabinose, and does so in a reversible manner. We believe it is a good alternative to existing growth control methods, especially for controlling co-culture, for several reasons. You can read more about our characterisation data on our results page.
→The system can easily be induced by L-arabinose, and switched off by d-glucose, both of which are non-toxic. .
→Gp2 is particularly good for regulating population size in co-cultures, as it only affects the size of one population.
Figure 2: Growth inhibition of Top10 cells by induction of Gp2 by arabinose
Figure 3: Recovery of growth by Top10 cells after induced araBAD operon is switched off by glucose-mediated catabolite repression.
