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Team:Evry/BXcom
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<h2>Overview</h2> | <h2>Overview</h2> | ||
| - | <p>Before engineering the plant hormonal system directly by the tadpole, we tested first if this hormone was inoffensive for the tadpole. We thought about how to deliver this hormone directly into the | + | <p>Before engineering the plant hormonal system directly by the tadpole, we tested first if this hormone was inoffensive for the tadpole. We thought about how to deliver this hormone directly into the embryos and the tadpoles. The first, and most obvious, decision was to inject directly into the embryos with microinjection, and, for the tadpoles to mix the MMR medium with auxin. The second, and most creative, was to use bacteria as a delivery machine in order to create a communication between two engineered organisms.</p> |
<h2>Steps</h2> | <h2>Steps</h2> | ||
| - | <p>Our idea was to use previous biobricks from Imperial College 2011 (BBa_K515100). Indeed, they managed to express in Escherichia coli the genes | + | <p>Our idea was to use previous biobricks from Imperial College 2011 (BBa_K515100). Indeed, they managed to express in <i>Escherichia coli</i> the genes encoding the IAA-producing pathway from <i>Pseudomonas savastanoi</i>. Besides, we constructed a plasmid with a reporter (mRFP) as a control to see how far the auxin production can go.</p> |
<center><img src="https://static.igem.org/mediawiki/2012/e/eb/Fig1.png"/></center><br> | <center><img src="https://static.igem.org/mediawiki/2012/e/eb/Fig1.png"/></center><br> | ||
<center><u>Figure 1: Delivery by bacteria engineered</u></center> | <center><u>Figure 1: Delivery by bacteria engineered</u></center> | ||
| - | <p>Once all the construction prepared/analyzed in DH5a bacteria, we prepared a mix with bacteria and MMR medium and LB medium. So in a 16 plates, wich each contains | + | <p>Once all the construction prepared/analyzed in DH5a bacteria, we prepared a mix with bacteria and MMR medium and LB medium. So in a 16-well plates, wich each contains three tadpoles we proceedeed as: <br> |
<center><img src="https://static.igem.org/mediawiki/2012/8/86/Rfpx.jpg"/></center><br> | <center><img src="https://static.igem.org/mediawiki/2012/8/86/Rfpx.jpg"/></center><br> | ||
<center><u>Figure 2: Fluorescence quantification</u></center> | <center><u>Figure 2: Fluorescence quantification</u></center> | ||
| - | The results show that, except for the control, all tadpoles are fluorescent. The Fig.2 shows also that the | + | 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 for 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.</p> |
Revision as of 13:35, 25 September 2012
Communication Bacteria<->Xenopus
Overview
Before engineering the plant hormonal system directly by the tadpole, we tested first if this hormone was inoffensive for the tadpole. We thought about how to deliver this hormone directly into the embryos and the tadpoles. The first, and most obvious, decision was to inject directly into the embryos with microinjection, and, for the tadpoles to mix the MMR medium with auxin. The second, and most creative, 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 see how far the auxin production can go.
Once all the construction prepared/analyzed in DH5a bacteria, we prepared a mix with bacteria and MMR medium and LB medium. So in a 16-well plates, wich each contains three tadpoles we proceedeed as:
