We first produced our gRNA plasmids in order to stop expression of targeted fluorescent proteins on our reporter plasmid.
How
Results
STEP 2: Create Reporter Plasmid
Why
How
Results
STEP 3: Promoter Library
Why
How
Results
STEP 4: Random Ligations
Why
How
Results
STEP 5: Freeze Drying & Revival
Why
For Key. coli to work as intended and not deteriorate we need need to things to occur:
The E. coli cells must be kept inactive so that nutrients is not depleted causing the transformed cells to die
The E. coli cells must be able to be activated after inactivation to allow the fluorescent genes to be expressed to give the key its unique fluorescent code which will allow access to the appliance.
How
To accomplish this, we chose to freeze dry the cells within the key. Click here for out protocol for freeze-drying cells.
Results
Figure 1: Graph of strong promoter 4 and weak promoter 1 transformed cells RFP fluorescence assay after freeze-drying revival two weeks and three weeks after samples were freeze dried.
In our results for freeze-dried cell revival, seen in Figure 1, we can show that storage temperatures do not have an effect on the revival of cells. For strong promoter 4 (SP4), at timepoint of 4 hours and 6 hours the storage temperature does seem to have a negative affect on the relative RFP fluorescence. However our sample size is very small meaning further assays would be needed to confirm this.
