Team:Czech Republic/Goals

Goals


Module 1 builds synthetic haploid strains with refactored mating loci that are conjugated to make a functional IOD. These strains have the wild-type mating phenotype and differentially express a reprogrammed signalling pathway in their diploid state proving the feasibility of the clone-free assembly concept.

Construct a set of reporter promoters for yeast cells
Characterize reporter promoters in all mating types
Design and materialized synthetic MATa and MATx strains
Build a synthetic diploid strain with a functional yeast pheromone pathway
Demonstrate the correct functionality of yeast pheromone pathway in synthetic diploids

Module 2 builds a set of orthogonal pheromones and receptors. These pheromone-receptor pairs enable specific localized signalling proving the feasibility of multichannel signal transmission underlying logic operations necessary for reliable diagnosis.

Construct a set of yeast plasmids with different mating pheromones and their receptors and contribute to the Registry with BioBricks
Verify the correct coupling of the receptors to the yeast pheromone mating pathway
Verify the correct expression and secretion of the different pheromones
Show the orthogonality of the used receptors and pheromones

Module 3 builds a set of location tags that recognize common tumor surface markers and agglutinate cell populations. Location tags displayed in the correct conformation strengthen cell-cell interactions to enable localization of signal transmission.

Express streptavidin, EpCAM, Anti-EpCAM scFv, c-Myc scFv and anti-HuA scFv on the surface of yeasts
Demonstrate the ability of our receptors to bind chosen markers
Monitor the dynamic binding of our receptors and their corresponding markers

Module 4 provides modeling and simulation support for other modules.

Develop a simulation environment CeCe to capture the complexity of cell-cell signal transmission
Design an IOD chemical reaction network model
Develop a schematic architecture for conceptual modeling of signal transmission networks
Design a two IOD signal transmission network suitable for the IOD band
Illustrate the robustness and efficiency of the IOD band design in CeCe simulations

Module 5 allows other modules to use microfluidics devices for their experiments.

Set of microfluidic devices fabricated by PDMS soft-lithography.
Characterization of signal transmission range between wildtype MATa and MATx Saccharomyces cerevisiae cells.
Dynamic characterisation of signal transmission between synthetic MATa and MATx Saccharomyces cerevisiae cells.
Comparison with mathematical model of signal transmission mechanism and estimation of the activation threshold for different cell concentrations.
Demonstrate yeast induced blood agglutination on-chip by human antigen A displayed on cell surface by Yeast Surface Display.