Team:Hannover/Results/Heavy Metals/Arabidopsis

From 2014.igem.org

(Difference between revisions)
Line 6: Line 6:
<body>
<body>
<h1><a href="https://2014.igem.org/Team:Hannover/Results_Project">Results</a> / Heavy metals / <i>Arabidopsis thaliana</i> T4MBP</h1>
<h1><a href="https://2014.igem.org/Team:Hannover/Results_Project">Results</a> / Heavy metals / <i>Arabidopsis thaliana</i> T4MBP</h1>
-
<h2>Labwork</h2><p class="text">Transformation of <i>A. thaliana</i> with our T4MBP:<ol><li><b></b>Exchange the original promoter enTCUP2 of the binary vector pORE_E3 <a href="http://www.ncbi.nlm.nih.gov/nuccore/AY562536">(AY562536.1)</a> into a 2x35s promoter using resctriction site based cloning.</li><li>We integrated our T4MBP in our modified pORE_E3_2x35S.</li><li>Application of our <a href="https://2014.igem.org/Team:Hannover/Protocols/Transformation/Arabidopsis#">floral dip </a>method.</li><li>After 6 weeks we harvested transgenic seeds from <i>A. thaliana</i> and plated these seeds on selection MSO-media.</li><li>At last we potted transformed plants.</li></ol></p>
+
<h2>Labwork</h2><p class="text">Transformation of <i>A. thaliana</i> with our T4MBP:<ol><li><b></b>Exchange the original promoter <a href="http://www.ncbi.nlm.nih.gov/nuccore/AY562536">enTCUP2</a> of the binary vector pORE_E3 <a href="http://www.ncbi.nlm.nih.gov/nuccore/AY562536">(AY562536.1)</a> into a 2x35s promoter using resctriction site based cloning.</li><li>We integrated our T4MBP in our modified pORE_E3_2x35S.</li><li>Application of our <a href="https://2014.igem.org/Team:Hannover/Protocols/Transformation/Arabidopsis#">floral dip </a>method.</li><li>After 6 weeks we harvested transgenic seeds from <i>A. thaliana</i> and plated these seeds on selection MSO-media.</li><li>At last we potted transformed plants.</li></ol></p>
<h2>Results</h2>
<h2>Results</h2>
Line 16: Line 16:
<br>
<br>
-
<p class="text">Below in figures 2 and 3 you can see the original vector pORE_E3 with an enTCUP3 promoter. For a better expression of our T4MBP protein we exchanged the enTCUP3 with the 2x35S promoter. Each step of this procedure is visualized in <a href="https://static.igem.org/mediawiki/2014/d/d1/Hannover_20141015_PORE_E3_2x35S%2BInsert_Anke_Fabian.dna_History.png">this histroy </a>. Furthermore this history includes the insertion of our T4MBP (there termed CDS for coding sequence). </p>
+
<p class="text">Below in figures 2 and 3 you can see the original vector pORE_E3 with an enTCUP2 promoter. For a better expression of our T4MBP protein we exchanged the enTCUP2 with the 2x35S promoter. Each step of this procedure is visualized in <a href="https://static.igem.org/mediawiki/2014/d/d1/Hannover_20141015_PORE_E3_2x35S%2BInsert_Anke_Fabian.dna_History.png">this histroy </a>. Furthermore this history includes the insertion of our T4MBP (there termed CDS for coding sequence). </p>
<center><table border="0">
<center><table border="0">

Revision as of 21:03, 16 October 2014

Results / Heavy metals / Arabidopsis thaliana T4MBP

Labwork

Transformation of A. thaliana with our T4MBP:

  1. Exchange the original promoter enTCUP2 of the binary vector pORE_E3 (AY562536.1) into a 2x35s promoter using resctriction site based cloning.
  2. We integrated our T4MBP in our modified pORE_E3_2x35S.
  3. Application of our floral dip method.
  4. After 6 weeks we harvested transgenic seeds from A. thaliana and plated these seeds on selection MSO-media.
  5. At last we potted transformed plants.

Results

The exchange of the promoter was checked by sequencing. We achieved to regenerate young A. thaliana after the transformation with our T4MBP. To receive a stabile F2 generation seeds of transformed plants would have to be harvested and grown. As a next step of this test series the plants would have been transfered on medium with heavy metals. In a following analysis it would be detected if these plants bind the heavy metal zinc, copper, cadmium and/or arsenic.

Fig. 1: Pictures showing young potentially transgenic A. thaliana before, during and after the transfer from medium to earth.


Below in figures 2 and 3 you can see the original vector pORE_E3 with an enTCUP2 promoter. For a better expression of our T4MBP protein we exchanged the enTCUP2 with the 2x35S promoter. Each step of this procedure is visualized in this histroy . Furthermore this history includes the insertion of our T4MBP (there termed CDS for coding sequence).

Fig. 2: Vector pORE_E3 with the original enTCUP2 promoter.

Fig. 3: Vector pORE-E3 with 2x35S promoter and our T4MBP which includes a sequence for expansin 4, cellulose-binding domain and domains for the binding of copper, arsenic, zinc, cadmium.