Our aim in this project was to create 3 gene assembly composite part as the genetic circuitry of PEred. We've had come to realize that 7 months seems less as opposed to colossal amount of wetlab we had in hand. Therefore, we instead shifted our focus on the characterization of the KdpF promoter and the behavior of individual genes in the presence of blood before we assemble all of these 3 elements together.
For this iGEM season our team is submitting the following part :
BBa_K3306002 is a composite biobrick part comprising of mutated version of wildtype KdpF promoter BBa_K1682004 upstream of GFP reporter BBa_E0040.
When cloned upstream of a reporter or any other protein coding gene, the part will cause expression of the protein in response to potassium ions. As per characterization by iGEM HKUST-Rice 2015, the dynamic range for that operation of the promoter is between 0 to 0.1 mM of K+ ions in external growth media of the chassis organism.
This data gave us a good reference as a starting point. But it was crucial to detect the activity of this promoter in the presence of blood/serum. PEred's success depends on its ability to sense external K+ ions from the menstrual blood and therefore the characterization data of our promoter repertoire in the presence of serum gave us some insight as to how reliable the promoter will be for the proper function of PEred's genetic circuity.
Having gained data regarding the activity of individual genes in the required substrate, our future plan include creating a composite part with KdpF promoter upstream of Laccase and OmpR234 and carrying out validation for this composite part.
