We could successfully clone our designed basic parts into the protein expression vector pBxC3H, show the expression of the proteins needed for our biosensor VbrK, VbrR and BlaR1. Additionally we were able to purifiy VbrK and BlaR1.
We have proven the ability of the cells to stay alive on the paper strip and their ability to react reliably to chemicals dissolved in water. BL21 cells, transformed with a plasmid containing a lac promotor and a β-galactosidase, were loaded on a strip. A blue product resulting from the reaction was observed in all samples except the negative control, with different concentrations of IPTG and X-Gal leading to gradually changing color intensities.
Figure: Paper strip test with different IPTG concentrations and 1 mg/ml X-Gal in water.
We managed to encapsulate live cells in alginate capsules, while retaining their ability react to the outside enviroment. Using BL21 cells with a similar plasmid as before, expressing GFP instead of a β-galactosidase, the bobas showed fluorescence in the presence of IPTG, with the negative control staying dark.
Different IPTG concentrations also lead to a difference in fluorescence intensity over time. This proves, that it is possible to quantify the amount of antibiotics in the water sample with the fluorescence intensity, if a standard series with known concentrations is made first.
Figure: Plots pixel intensity over time.
We created a functioning systems viable for whole-cell biosensors. All that is left to do to finalize BLISS is to implement the plasmid containing the antibiotic receptors. Overall these systems can be applied in tandem with any whole cell biosensor reacting to liquid samples, making them extremely useful in reseach and enviromental studies.