Team:SSTi-SZGD/Description
SSTi-SZGD
Pesticide Residue Sweeper--Why We Need it?
The level of pesticide usage in China is 2.5 times above the world average. In June 2016, the total output of chemical pesticides in China was around 3.347 million tons, up by 7.17% compared with the same period of 2015. The deterioration effects of long-term and overdose usage of pesticide in soil include soil erosion, soil contamination, water pollution, organic pollution, reducing biodiversity, etc.
A range of chemical and physical methods have been applied to degrade pesticide residues. The shortfalls of these methods include high in cost, harmful to non-target organisms, food, plants and soils, and likely to cause secondary pollution. We intend to develop a microbial degradation method with low toxicity, low cost and high efficiency to serve a positive role in maintaining ecological balance.
What we will do?
SSTi-iGEM combined optogenetics and biotechnology, using genetically modified E.coli as a carrier, to improve the microbial method for pesticide residue degradation in soil. We used a light-regulated gene expression system (LightOFF) that efficiently over-express heterogenous hydrolases which degrade insecticide organophosphorus or fungicide carbendazim. This application can result to two forms of products: enzyme and whole cell products. In addition, we propose to develop an automatic praying system that facilitates the application of whole cell products in real-life scenario.
How to do it?----Project Design
Our LightOFF expression system constitutes a fusion protein of LexA repressor from E. coli SOS regulon, and a blue light sensor (VVD) from Neurospora crassa. Light irradiation causes conformational change of VVD and subsequent dimerization of the fusion protein. The activated dimer thus binds its cognate operator sequence and represses the promoter activity. We first tested its function and induction efficiency.
There are a few identified gene encoding hydrolases to detoxifiy organophosphate and carbendazim pesticides. After a PubMed and iGEM search, we came up with two candidate genes: opd A and mheI. The former has been used in other iGEM projects before and proven working, the latter was conserved in many bacteria species that use carbendazim as a carbon source. We also added a signal peptide from Tat translocase of E. coli to help with protein exportation.
We considered the risk of releasing GMOs to the natural environment. So we chose to inclulde a suicide gene in the system. Supernova, an engineered genetically-encoded photosensitizer, containing chromphores that generate reactive oxygen species (ROS) upon illumination, and can be used to promote apoptosis in prokaryotic cells. When combine with LightOFF system, darkness induces supernova expression, while light irradiation triggers the release of ROS to promote cell death.
By combining the above three elements together, we could efficiently produce enzyme (hydrolase) products or construct a live biocatalyst with applications in farms, orchards and gardens. We mainly focused on the product design, target customers, and cost analysis to identify markets. In addition, we developed an automatic spraying device to be used with live biocatalysts.
