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Team:HokkaidoU Japan/Shuffling Kit/Examples
From 2013.igem.org
Maestro E.coli
Optimization Kit
Demonstrations for Usecase Example
We will show some interesting demonstrations of our kits, Promoter Selector and RBS Selector!
Promoter Selector
Let's select the best promoter for Kanamycin resistance by Promoter Selector.
For a demonstration we decided to optimize the expression of Kanamycin resistance. Changing the concentration of Kanamycin in agar plate, it is estimated that different promoter will be chosen by our Promoter Selector (fig.1).
If the concentration of Kanamycin was high, the colony with strong promoter will survive. Therefore, only one or two colors of colonies would appear. If the concentration of Kanamycin was low, colonies with weak promoters will be able to survive. This way many colors of colonies would appear (fig.2).
Conclusion
Our Promoter Selector seemed to work. We got several colors colonies on Kanamycin plates. This means that Kanamycin Resistance gene was successfully ligated. We need more than 24 hrs to identify the colors.
RBS Selector
Let’s create all combinations by two reporter genes and make various colors on one plate!
The RBS Selector we made, can randomize the strength of RBSs in the operon. For a demonstration, we decided to create all combinations by two genes; mRFP1 (BBa_E1010) and LacZα (BBa_I732006) (fig.4). LacZα makes the colony blue. mRFP1 makes the colony red.
Conclusion
We can say that our RBS Selector worked!! The RBSs uptsream 2 genes were randomized and they had many levels of expressions.
