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From 2012.igem.org

Communication Bacteria<->Xenopus

Overview

Engineering Xenopus embryos with AID system raised the question of how will we deliver auxin to the embryonic cells ? One idea was to use bacteria as a delivery machine in order to create a communication between two engineered organisms.

Steps

Our idea was to use previous biobricks from Imperial College 2011 BBa_K515100. Indeed, they managed to express in Escherichia coli the genes encoding the IAA-producing pathway from Pseudomonas savastanoi. Besides, we constructed a plasmid with a reporter (mRFP) as a control to monitor the auxin production.

Figure 1: Enginereed bacteria delivery to tadpole

Embryos were placed in medium containing MMR and DH5a bacteria with either BBa_K515100 or a reporter (mRFP), as shown on below

Figure 2: Fluorescence observations with different Bacteria concentrations



Figure 3: Fluorescence observations by day



Figure 4: Proof of principle The results show that, except for the control, all tadpoles are fluorescent. The Fig.2 shows also that the fluorescence occurs mainly in the intestinal tract. The death rate during the experiments is close to 0%.
We've performed the same using the Imperial College 2011 plasmid; the tadpoles didn't show any sign of auxin intolerance.

Conclusion

Our results clearly show fluorescence in the tadpole, especially in the intestine. Thus, we demonstrate that it was possible to use bacteria in a way to deliver some specific molecules to the Xenopus. For our project, using the bacteria as an auxin factory represents a good alternative to the production devices 1 and 2, only if the auxin can join the circulatory system .