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Cambridge Team
You got us under the microscope

As a part of testing our bistable switch, we had to screen E. coli bacteria to detect fluorescence from GFP and RFP expression. Apart from the methods used in the lab, we also collaborated with the Cambridge iGEM team who designed the OpenScope microscope. (fig1) It is a 3D-printed brightfield and fluorescence imaging microscope that has the potential to be a low-cost tool accessible to anyone interested in microscopy. We therefore sent samples of bacteria expressing these proteins that they could use to test their microscope. As an outcome of this collaboration, we were also able to confirm some of our own results.

Fig 1: The OpenScope microscope designed by the Cambrigde team as seen from the side.

The samples we sent for imaging were:
• DH5α –no plasmid, no antibiotic resistance, no fluorescence
• DH5α - K1725103, Kanamycin resistance, GFP and RFP fluorescence
• DH5α - K1725043 and K1725103, Kanamycin and Chloramphenicol resistance, RFP fluorescence
• DH5α - K1725063 and K1725103, Kanamycin and Chloramphenicol resistance, GFP fluorescence
• DH5α - K1725100 and K1725103, Kanamycin and Chloramphenicol resistance, GFP fluorescence with a few colonies expressing RFP

The results that we received from the Cambridge team confirmed the presence of GFP in our cells agreeing with our results. (fig 2) Their microscope failed to detect RFP, but its expression was also confirmed by using a commercial microscope. It should also be mentioned that the brightness intensity of the fluorescent beads initially used is much greater than this of our fluorescent bacteria that are much smaller in size and dispersed on the plate. This can be seen in the Cambridge wiki page:

Fig 2: a) This image shows the negative control where there is no plasmid in the DH5α cells, and therefore there is no fluorescence.
b) The second image shows clustered DH5a cells with K1725063 and K1725103 plasmids. GFP is expressed in this case.


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