Safety

The design of our project required careful consideration as to the impact it might have on the environment. Our project would ideally be able to be released into a water system that was contaminated by high levels of nitrates. In its completed form it would be able to fully reduce the nitrates to nitrogen gas. With this in mind we needed to ensure that we didn’t transform a nitrate problem into an E. coli problem by introducing a strain of E. coli that would grow rampant.

In order to prevent the E. coli from growing excessively, design and incorporation of a kill switch was a large part of our project. We wanted to create a kill switch that would allow our strain to grow successfully when there are high levels of nitrate ions for it to reduce, but that prevents it from growing when nitrate levels are low and the strain is no longer necessary. After consideration of several methods to achieve this desired effect, we settled on complementing an auxotrophic strain of E. coli with a gene that would only be expressed under high nitrate conditions. To accomplish this we looked to previous iGEM teams for inspiration and quickly discovered the Edinburugh 2009 team’s PyeaR BioBrick that was shown to be a nitrate sensitive promoter for E. coli. We believed this promoter could be used with a serA gene to complement a ΔserA strain of E. coli that one of our advisers had previously worked with.

This work led to us designing and constructing the BioBrick BBa_K2086002. Preliminary testing done by one of our advisors showed that it is possible for this BioBrick to act as a successful kill-switch.