Difference between revisions of "Team:Vilnius-Lithuania/Demonstrate"
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<a href="https://2019.igem.org/Team:Vilnius-Lithuania/Model" class="sub">Overview</a> | <a href="https://2019.igem.org/Team:Vilnius-Lithuania/Model" class="sub">Overview</a> | ||
<a href="https://2019.igem.org/Team:Vilnius-Lithuania/Degradation_Rate_Control_Model" class="sub">Post-translational regulation</a> | <a href="https://2019.igem.org/Team:Vilnius-Lithuania/Degradation_Rate_Control_Model" class="sub">Post-translational regulation</a> | ||
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<a href="https://2019.igem.org/Team:Vilnius-Lithuania/Transcriptional_Regulation_CarH" class="sub">Transcriptional regulation (CarH)</a> | <a href="https://2019.igem.org/Team:Vilnius-Lithuania/Transcriptional_Regulation_CarH" class="sub">Transcriptional regulation (CarH)</a> | ||
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<a href="https://2019.igem.org/Team:Vilnius-Lithuania/Software">Software</a> | <a href="https://2019.igem.org/Team:Vilnius-Lithuania/Software">Software</a> | ||
<a href="#">Human Practices</a> | <a href="#">Human Practices</a> | ||
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<a href="https://2019.igem.org/Team:Vilnius-Lithuania/Degradation_Rate_Control_Model">Post-translational regulation</a> | <a href="https://2019.igem.org/Team:Vilnius-Lithuania/Degradation_Rate_Control_Model">Post-translational regulation</a> | ||
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<a href="https://2019.igem.org/Team:Vilnius-Lithuania/Transcriptional_Regulation_CarH">Transcriptional regulation (CarH)</a> | <a href="https://2019.igem.org/Team:Vilnius-Lithuania/Transcriptional_Regulation_CarH">Transcriptional regulation (CarH)</a> | ||
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| − | + | <p>The proof of Colight consist of several critical steps. </p><br> | |
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| − | + | <h4 class="page-heading">1.The proof that CarH works as light inducible repressor</h4><br> | |
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| − | + | <img src="https://2019.igem.org/wiki/images/c/c6/T--Vilnius-Lithuania--Trans-Carh1.png" style="width:100%"><br><br> | |
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| − | + | <p>The experiment shows that CarH expressing cells grown in the dark for 12 hours have a substantially reduced level of fluorescence. That is because CarH represses its promoter under dark conditions and the promoter is induced under light.</p><br> | |
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| + | <h4 class="page-heading">2.The proof of light inducible protein degradation by mf-Lon</h4><br> | ||
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| + | <img src="https://2019.igem.org/wiki/images/e/e3/T--Vilnius-Lithuania--Degradation-2.png" style="width:100%"><br><br> | ||
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| + | <p>In this graph we see that a protein of a construct number 160 has its protein level reduced after induction by light. That is because after induction asLOV change its conformation, and degradation tag becomes accessible to mf-Lon protease which degrades sfGFP.</p><br> | ||
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| + | <h4 class="page-heading">3.The proof of light modular plasmid copy number regulation</h4><br> | ||
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| + | <img src="https://2019.igem.org/wiki/images/a/a1/T--Vilnius-Lithuania--copy2.png" style="width:100%"><br><br> | ||
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| + | <p>In this graph we show that illuminated cells, compared to uniluminated, have their plasmid copy number increased (grown until 16 hours). And when the light is turned off for both bacteria culture samples, the plasmid copy number bacteria is equalized.</p><br> | ||
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Latest revision as of 03:46, 22 October 2019
Demonstration
The proof of Colight consist of several critical steps.
1.The proof that CarH works as light inducible repressor
The experiment shows that CarH expressing cells grown in the dark for 12 hours have a substantially reduced level of fluorescence. That is because CarH represses its promoter under dark conditions and the promoter is induced under light.
2.The proof of light inducible protein degradation by mf-Lon
In this graph we see that a protein of a construct number 160 has its protein level reduced after induction by light. That is because after induction asLOV change its conformation, and degradation tag becomes accessible to mf-Lon protease which degrades sfGFP.
3.The proof of light modular plasmid copy number regulation
In this graph we show that illuminated cells, compared to uniluminated, have their plasmid copy number increased (grown until 16 hours). And when the light is turned off for both bacteria culture samples, the plasmid copy number bacteria is equalized.