Team:Hong Kong JSS/Engineering
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Hong Kong JSS
Due to COVID-19, we have very limited access to our school laboratory. As a result, we have joined the two-phase project such that work can be distributed across two years. In order to demonstrate engineering success, we made effort to follow the engineering design cycle:
Design → Build → Test → Learn → Design
Since this is phase 1 of our project, more effort will be spent on design and modeling.
Design
Current methods to reduce aflatoxin often involve the use of heating, gamma rays or sorbent additives, which will reduce the nutrient level in food. Our project aimed to provide a better alternative in fighting aflatoxin contamination. The design of our system involves the production of tvLac and FDR-A - two enzymes capable of degrading aflatoxins, in genetically modified E.coli. (details of tvLac and FDR-A can be found in basic parts page: https://2021.igem.org/Team:Hong_Kong_JSS/Basic_Part)
Build
In our project, we planned to produce 2 products. One is a “Detox-spray” using purified tvLac and FDR-A extracted from GM E. coli. Another will be a probiotic E. coli strain “AflaStop”, capable of competing with the aflatoxin-producing fungus and secret tvLac and FDR-A to degrade any aflatoxins in its surrounding (for details: https://2021.igem.org/Team:Hong_Kong_JSS/Proof_Of_Concept).
We planned in phase 2 to build the following composite parts for protein purification and secretory expression of our tvLac and FDR-A:
tvLac and FDR-A are tagged with a 6-His Tag (CATCATCATCATCATCAT) for protein purification.
In the secretory constructs, either signal peptide DsbAss (BBa_K3746005) or NSP4 (BBa_K3746004) were added to the N-terminal for the protein secretion.
For details of each bio brick used in the above composite parts, please refer to our parts registry. The list of all parts can be found at: https://2021.igem.org/Team:Hong_Kong_JSS/Parts
We planned in phase 2 to build the following composite parts for protein purification and secretory expression of our tvLac and FDR-A:
tvLac and FDR-A are tagged with a 6-His Tag (CATCATCATCATCATCAT) for protein purification.
In the secretory constructs, either signal peptide DsbAss (BBa_K3746005) or NSP4 (BBa_K3746004) were added to the N-terminal for the protein secretion.
For details of each bio brick used in the above composite parts, please refer to our parts registry. The list of all parts can be found at: https://2021.igem.org/Team:Hong_Kong_JSS/Parts
Test
We had performed preliminary tests to verify our concept. The result of our test confirmed the following:
For details of our testing procedures and planned experiments for phase 2, please visit: https://2021.igem.org/Team:Hong_Kong_JSS/Experiments
For the results of these tests, please visit: https://2021.igem.org/Team:Hong_Kong_JSS/Results
- We can cultivate Aspergillus flavus in our lab conditions.
- We can extract and detect dilute aflatoxins produced by Aspergillus flavus.
- The extracted dilute aflatoxins can be degraded by Laccase, we can detect the disappearance of aflatoxins after digestion by Laccase.
For details of our testing procedures and planned experiments for phase 2, please visit: https://2021.igem.org/Team:Hong_Kong_JSS/Experiments
For the results of these tests, please visit: https://2021.igem.org/Team:Hong_Kong_JSS/Results
