What is Nitrate Neutralizer?
UMD’s 2022 iGEM project uses synthetic biology to address nitrate pollution, which causes severe damage to our Chesapeake Bay watershed. When fertilizers containing nitrogen are deposited into the waterways, they cause algae to overgrow, which blocks sunlight from bay grasses and depletes the water of oxygen when bacteria break the algae down. This process (eutrophication) can lead to dead zones, which are areas that cannot sustain aquatic life because they lack oxygen.
Additionally, the process of creating nitrogen based fertilizers leads to significant carbon emissions and contributes to climate change, which also affects our bay ecosystem.
In order to maintain the health of the Chesapeake Bay, significant work must be done to mitigate nitrate levels in the water and develop a system that reuses nitrogen from fertilizer to limit carbon emissions.
Our project tackles this issue with three objectives:
- Genetically engineer an organism to take in nitrogen
- Include a mechanism in the genetic system that allows the nitrogen to be reused
- Encapsulate the organisms in a bioreactor system
Our project would remove nitrates from the water using genetically engineered Escherichia coli. The E.coli in our system, once transformed with the genes cyanophycin synthetase (CphA) and cyanophycinase (CphB), would be able to take in nitrates from the water and store them as nitrogen granules called cyanophycin granule polypeptide (CGP). The CGP, which is synthesized by cyanophycin synthetase from the CphA gene, can then be converted into nitrogen rich amino acids with the help of cyanophycinase from the CphB gene.
In order to transform the E.coli with the CphA and CphB genes, the following plasmids were designed using iGEM biobricks.
Plasmid A was designed with a constitutive Weak Anderson promoter to allow CphA to be transcribed consistently, while plasmid Plasmid B was designed with an inducible plac promoter that would only transcribe CphB in the presence of lactose. These plasmids were combined using Golden Gate Assembly and transformed into E. coli.
Once transformed with the plasmid, the E.coli were encapsulated within alginate beads to prevent them from escaping into bay water. These beads were then placed in a bioreactor, which feeds the bacteria bay water with nitrates to test their nitrogen storage capabilities.
This system, with more research, could potentially help farmers filter nearby waterways of nitrogen runoff from fields, while also allowing the nitrogen to be reused for fertilizer after harvesting the alginate beads.