Team:NYMU-Taipei/Design
Design
Begin with the End in Mind
1. Set Our Goal
Develop non-invasive odorant biosensors for early screening of human diseases.
2. Focus Specific Aim
Detect heptanal which can be used as a breath screening marker for lung cancer and tuberculosis disease.
3. Develop The Strategy
Use surface-displayed and intracellular expressed proteins to test the binding specificity and sensitivity.
4. Identify The Needs
- Gene of olfactory receptor protein
- Expressed olfactory receptor protein on cell surface
- Expressed olfactory receptor protein in cells
- Control proteins that do not bind to heptanal
- Immobilization method to fix proteins to cellulose chromatography paper
- Binding method for binding heptanal onto immobilized proteins
- Detection method for measuring the binding specificity and sensitivity
- Prototype device for collecting volatile biomarkers for breath screening
5. Develop The Scope
To prove our DiseaScent concept, we decide to express mOR103-15, an olfactory receptor protein that binds to heptanal, in E. coli cells. Also, fluorescent proteins with different multi-meric forms (monomer. Dimer, tetramer) are cloned and expressed side-by-side to use as negative controls.
6. Identify The Team
- Gene Cloning Team
- Protein Expression Team
- Protein Immobilization Team
- Protein-heptanal Binding Detection Team
- Team for Prototype Device for Collecting Volatile Biomarkers for Breath Screening
7. Define The Process
- Decide and obtain the needed gene of olfactory receptor protein
- Decide and obtain the needed vector for expressing olfactory receptor protein
- Surface display
- Intracellular expression
- Decide and obtain the needed vector for expressing various fluorescent proteins
- mCherry
- eforCP
- AmpilCP, Blue
- sfGFP
- Design cloning strategy and design PCR primers
- Perform PCR experiments to get both the insert gene of olfactory receptor protein and the E. coli expression vector
- Perform PCR experiments to get both the insert genes of various fluorescent proteins and the E. coli expression vector
- Gel extraction and ligation, transformation experiments
- Identification of correctly cloned gene of olfactory receptor protein
- Surface display
- Intracellular expression
- Identification of correctly cloned genes of various fluorescent proteins
- Intracellular expression
- Perform protein expression induction and identification
- Perform protein immobilization experiments
- Decide the appropriate cellulose chromatography paper to use for protein immobilization
- Treat the cellulose chromatography paper for protein immobilization capability
- Test the treated cellulose chromatography paper for protein immobilization
- Adjust and optimize the binding efficiency
- Measure binding specificity and sensitivity
- Design prototype device for collecting volatile biomarkers for breath screening
- Review current methods for collecting volatile biomarkers
- Define our simple-to-carry and simple-to-use volatile biomarkers collecting prototype
- Design our simple-to-carry and simple-to-use volatile biomarkers collecting prototype
- Combine our immobilized proteins on paper with the volatile biomarkers collecting prototype