Team:Toulouse/Result/experimental-results

From 2014.igem.org

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<p class="legend">Figure 18: Microscopic view of bead surfaces coated with chitin</p>
<p class="legend">Figure 18: Microscopic view of bead surfaces coated with chitin</p>
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<p class="texte">Using ImageJ software, we are able to create 3D pictures and movies of those comments. </br></p>
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<p class="texte">Using ImageJ software, we are able to create 3D pictures and movies of those comments.</br></p>
<center><img src="http://2014.igem.org/wiki/images/5/53/Photo_billes_microscopie.png" width="45%"><iframe width="380" height="315" src="//www.youtube.com/embed/ztIHIKQr3g0" frameborder="0" allowfullscreen></iframe></center>
<center><img src="http://2014.igem.org/wiki/images/5/53/Photo_billes_microscopie.png" width="45%"><iframe width="380" height="315" src="//www.youtube.com/embed/ztIHIKQr3g0" frameborder="0" allowfullscreen></iframe></center>
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<p class="texte">The first tests were accomplished with commercial GAFP-1 and D4E1 peptides at different concentrations (12.5µM, 25µM, 100µM). These tests were performed on different fungal strains sharing the same phylum with <i>Ceratocystis Platani</i>.
<p class="texte">The first tests were accomplished with commercial GAFP-1 and D4E1 peptides at different concentrations (12.5µM, 25µM, 100µM). These tests were performed on different fungal strains sharing the same phylum with <i>Ceratocystis Platani</i>.
As <i>Ceratocystis Platani</i> is pathogenic, we could not perform tests directly with this fungus.</br>
As <i>Ceratocystis Platani</i> is pathogenic, we could not perform tests directly with this fungus.</br>
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After several days at 30°C, the PDA (Potato Dextrose Agar) plates couvered with fungus and commercial peptides were analyzed.</p></p>
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After several days at 30°C, the PDA (Potato Dextrose Agar) plates covered with fungus and commercial peptides were analyzed.</p></p>
<p class="texte">An inhibition halo was noticeable with commercial D4E1 peptide at 100µM on <i>Aspergillus brasiliensis</i>. Less bright halos were also present with lower concentrations. Concerning commercial GAFP-1, we did not notice any effect in the tested conditions.As positive control, a well-known chemical fungicide was used: the Copper Sulfate. The inhibition of the fungal growth was complete at 20mg/ml, and at 10mg/ml a darker halo appeared around the pad filled with Copper Sulfate as we can see on the figure below.  This corresponds to a sporing halo in response to the stress generated by the fungicide.
<p class="texte">An inhibition halo was noticeable with commercial D4E1 peptide at 100µM on <i>Aspergillus brasiliensis</i>. Less bright halos were also present with lower concentrations. Concerning commercial GAFP-1, we did not notice any effect in the tested conditions.As positive control, a well-known chemical fungicide was used: the Copper Sulfate. The inhibition of the fungal growth was complete at 20mg/ml, and at 10mg/ml a darker halo appeared around the pad filled with Copper Sulfate as we can see on the figure below.  This corresponds to a sporing halo in response to the stress generated by the fungicide.
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The choice of our chassis appears to be optimal as we noted that wild type <i>Bacillus subtilis</i> disturbs the hyphae growth of the fungi. Some strains of <i>Bacillus subtilis</i> (qst 713) are already used as Biofungicides for use on several minor crops to treat a variety of plant diseases and fungal pathogens.</br>
The choice of our chassis appears to be optimal as we noted that wild type <i>Bacillus subtilis</i> disturbs the hyphae growth of the fungi. Some strains of <i>Bacillus subtilis</i> (qst 713) are already used as Biofungicides for use on several minor crops to treat a variety of plant diseases and fungal pathogens.</br>
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After this set of experiments, the strains expressing D4E1 and expressing GAFP-1 + D4E1 have shown to be the best candidates to play a major role in the fight against fungal diseases such as Canker stain. Keeping in mind our objective, <b> we decided to tests these strains in model plants </b> : <i>Nicotiana benthamiana</i> and <i>Arabidopsis thaliana</i>.</br> These tests were performed in the National Institute for the Agronomic Research by experts in this domain.  
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After this set of experiments, the strains expressing D4E1 and expressing GAFP-1 + D4E1 have shown to be the best candidates to play a major role in the fight against fungal diseases such as Canker stain. Keeping in mind our objective, <b> we decided to tests these strains in model plants</b>: <i>Nicotiana benthamiana</i> and <i>Arabidopsis thaliana</i>.</br>
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These tests were performed in the National Institute for the Agronomic Research by experts in this domain.  
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<p class="title2">3. <i> In planta </i> tests with SubtiTree<p/>
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<p class="title2">3. <i>In planta</i> tests with SubtiTree</p>
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<img style="width:215px;" img src="http://parts.igem.org/wiki/images/a/af/In_planta.jpg" style="margin-top:5px" />
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<img style="width:215px;" img src="http://parts.igem.org/wiki/images/a/af/In_planta.jpg" style="margin-top:5px"/>
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<p class="legend"> Figure 23: Injection of SubtiTree in a model plant </p>
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<p class="legend"> Figure 23: Injection of SubtiTree in a model plant</p>
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Revision as of 21:48, 15 October 2014