Difference between revisions of "Team:TU Darmstadt/Demonstrate"
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<a href="https://2017.igem.org/Team:TU_Darmstadt/project/chemistry#results">See more Details</a> | <a href="https://2017.igem.org/Team:TU_Darmstadt/project/chemistry#results">See more Details</a> | ||
</p></div> | </p></div> | ||
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| − | <img src="https://static.igem.org/mediawiki/2017/ | + | |
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| − | </figcaption></ | + | <figure><div style="display: flex; align-items:center;"> |
| − | </div> | + | <div style="display: flex; flex-direction:column; margin-right: 20px;"> |
| + | <img src="https://static.igem.org/mediawiki/2017/8/8d/T--TU_Darmstadt--8mMolPeptide.png" style="float: left; width:100%; margin-right:0.5%; margin-bottom: 0.5em;"><figcaption><b>Figure 4. ChiTUcare in unactivated state. </b></figcaption></div> | ||
| + | <div style="display: flex; flex-direction:column; margin-right: 20px;"> | ||
| + | <img src="https://static.igem.org/mediawiki/2017/1/1d/T--TU_Darmstadt--tomi1.png" style="float: left; width:100%; margin-right:0%; margin-bottom: 0.5em;"> | ||
| + | <figcaption><b>Figure 5. left: Fluormetric measurement of the peptide cleavage via proteases. right: cuvette filled this the product under UV-light.</b></figcaption></div></div> | ||
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</section> | </section> | ||
| − | <section id=" | + | <section id="Proof"><div class="container"> |
<h2>ChiTUcare</h2> | <h2>ChiTUcare</h2> | ||
<div class="post-it" style="margin-bottom:30px; display: flex; align-items:center;"> | <div class="post-it" style="margin-bottom:30px; display: flex; align-items:center;"> | ||
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<figure><div style="display: flex; align-items:center;"> | <figure><div style="display: flex; align-items:center;"> | ||
<div style="display: flex; flex-direction:column; margin-right: 20px;"> | <div style="display: flex; flex-direction:column; margin-right: 20px;"> | ||
| − | <img src="https://static.igem.org/mediawiki/2017/7/71/T--TU_Darmstadt--unaktiviert.jpg" style="float: left; width:100%; margin-right:0.5%; margin-bottom: 0.5em;"><figcaption><b>Figure | + | <img src="https://static.igem.org/mediawiki/2017/7/71/T--TU_Darmstadt--unaktiviert.jpg" style="float: left; width:100%; margin-right:0.5%; margin-bottom: 0.5em;"><figcaption><b>Figure 6. ChiTUcare in unactivated state. </b></figcaption></div> |
<div style="display: flex; flex-direction:column; margin-right: 20px;"> | <div style="display: flex; flex-direction:column; margin-right: 20px;"> | ||
<img src="https://static.igem.org/mediawiki/2017/5/59/T--TU_Darmstadt--aktiviert.jpg" style="float: left; width:100%; margin-right:0%; margin-bottom: 0.5em;"> | <img src="https://static.igem.org/mediawiki/2017/5/59/T--TU_Darmstadt--aktiviert.jpg" style="float: left; width:100%; margin-right:0%; margin-bottom: 0.5em;"> | ||
| − | <figcaption><b>Figure | + | <figcaption><b>Figure 7. ChiTUcare activated by protease.</b></figcaption></div></div> |
</figure> | </figure> | ||
<div style="margin-top: 40px;"> | <div style="margin-top: 40px;"> | ||
Revision as of 16:32, 30 October 2017
ChiTUcare
Proof of Concept
Here we want to give you a short tour about what we achieved in the project. We show our results and give a view on the resulting product, an protease-sensing hydrogel.
Chitin Synthase NodC
We expressed the chitin synthase NodC in E. coli Top10 successfully. The protein was tagged with a His-tag and purified via an ÄKTA system. To verify the expression and purification a SDS-PAGE was done.
The functionality of the NodC enzyme was verified by performing the UDP-Glo™ Glycosyltransferase Assay. The evaluation of the assay shows that the NodC enzyme converts the UDP-GlcNAc to free UPD and a growing oligo-GlcNAc-chain. The free UDP is converted to ATP, which acts as a substrate for a luciferase reaction and creates luminescence. So the assay and the increasing luminescence depending on increasing enzyme concentrations shows that the NodC enzyme can create chitin oligomers.
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Under Construction
Hydrogel
We manufactured several hydrogels containing non-toxic and cost-effective gelling agents to form optimal wound dressings. Our hydrogels could be formed with basic laboratory equipment at any shape and could easily be adjusted to the affected tissues for optimal wound healing.
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Chemistry
We manufactured a protease-sensing chitosan derivative. For the verifaction of its functionality it has been measured via flourimeter. It showed a peak at 390 nm before the protease was added. After the cleavage with the protease a shift of this peak to the wavelength of 450 nm was observed.
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ChiTUcare
To show that our project works, we manufactured a chitosan hydrogel with a fluorophore coupled to it.
Upon addition of a protease solution to our ChiTUcare prototype, the hydrogel starts to glow under UV light within 7 minutes (the video speed was increased).
This proves our concept of a protease sensing hydrogel on the basis of chitosan.
As an easy and cheap to manufacture prototype, we provide an eco-friendly and adjustable system to nurture wound-healing with an antibacterial, anti-viral wound dressing, that works as a sensor for bacterial infection by emitting a blue light within minutes upon irradiation of non-hazardous UV light.
