Difference between revisions of "Team:Toronto/Composite Part"
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− | <h2 id=" | + | <h2 id="tode-and-todf-plasmid-design">TodE and TodF Plasmid Design</h2> |
− | < | + | <p>Pseudomonas Putida F1 species degrade toluene with the help of tod operon. Toluene dioxygenase and toluene cis-dihydrodiol dehydroxygenase transcribed from transcribed from todC1C2BA and todD genes respectively facilitate break down of toluene to 3-methylcatechol. Gerben J. Zylstra et al successfully constructed the plasmid pTDG602 containing todC1C2BAD genes. To achieve complete degradation of toluene down to carbon dioxide and water, we focused on designing a plasmid containing todE and todF genes. After cotransformation of DH10B cells with pTDG602 and our plasmid, E.Coli theoretically should contain all the genes required to achieve complete degradation.</p> |
− | <p>In order to | + | <p><img src="https://github.com/igemuoftATG/wiki2015/blob/master/images/Pathway.png" alt="alt text"></p> |
− | + | <p>In order to achieve the best protein production efficiency, we codon optimized our todE and todF sequences. We used Salis RBS calculator to optimize translation efficiency. We used a tunable promoter to determine optimum transcription for DH10B cells.</p> | |
− | this | + | <p><img src="https://github.com/igemuoftATG/wiki2015/blob/master/images/biobrick%20plasmid.png" alt="alt text"></p> |
− | + | <p>We also used BioBricks in the registry to design plasmids with different efficiency by using RDP kit as shown below. In this process, we used promoter BBa_J23102 and RBS BBa_B0030.</p> | |
− | + | <p><img src="file://localhost/Users/joannadowdell/Downloads/TodEplasmid.webp" alt="alt text"></p> | |
− | + | <p><img src="https://github.com/igemuoftATG/wiki2015/blob/master/images/TodFplasmid.png" alt="alt text"></p> | |
− | <p> | + | <p>Note: Links to iGem registry |
− | + | BBa_J23102: <a href="http://parts.igem.org/Part:BBa_J23102">http://parts.igem.org/Part:BBa_J23102</a> | |
− | <a href="http://parts.igem.org/ | + | BBa_0030: <a href="http://parts.igem.org/Part:BBa_B0030">http://parts.igem.org/Part:BBa_B0030</a></p> |
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Revision as of 23:03, 18 September 2015
TodE and TodF Plasmid Design
Pseudomonas Putida F1 species degrade toluene with the help of tod operon. Toluene dioxygenase and toluene cis-dihydrodiol dehydroxygenase transcribed from transcribed from todC1C2BA and todD genes respectively facilitate break down of toluene to 3-methylcatechol. Gerben J. Zylstra et al successfully constructed the plasmid pTDG602 containing todC1C2BAD genes. To achieve complete degradation of toluene down to carbon dioxide and water, we focused on designing a plasmid containing todE and todF genes. After cotransformation of DH10B cells with pTDG602 and our plasmid, E.Coli theoretically should contain all the genes required to achieve complete degradation.
In order to achieve the best protein production efficiency, we codon optimized our todE and todF sequences. We used Salis RBS calculator to optimize translation efficiency. We used a tunable promoter to determine optimum transcription for DH10B cells.
We also used BioBricks in the registry to design plasmids with different efficiency by using RDP kit as shown below. In this process, we used promoter BBa_J23102 and RBS BBa_B0030.
Note: Links to iGem registry BBa_J23102: http://parts.igem.org/Part:BBa_J23102 BBa_0030: http://parts.igem.org/Part:BBa_B0030