Difference between revisions of "Team:Tuebingen/Description"
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<rect x="1000" y="120" rx="20" ry="20" width="800" height="510" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | <rect x="1000" y="120" rx="20" ry="20" width="800" height="510" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | ||
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− | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size: | + | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size:46px;font-weight:bold;color:black;">The expression of the Dronpa-Cre-Dronpa construct is under control of a promotor, that is influenced by a sensor. |
− | Any | + | Any biosensor that is able to positively regulate a promotor can be used in our setting.</p> |
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<rect x="10" y="10" rx="20" ry="20" width="1100" height="320" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | <rect x="10" y="10" rx="20" ry="20" width="1100" height="320" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | ||
<foreignObject x="10" y="10" width="1100" height="320" > | <foreignObject x="10" y="10" width="1100" height="320" > | ||
− | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size:52px;font-weight:bold;color:black;">After expression the Dronpa-Cre- | + | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size:52px;font-weight:bold;color:black;">After expression the Dronpa-Cre-Dronpa construct is constituted of fluorescent Dronpa domains that attach to each other and thereby inhibit the Cre domain sterically.</p> |
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<rect x="200" y="780" rx="20" ry="20" width="1100" height="300" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | <rect x="200" y="780" rx="20" ry="20" width="1100" height="300" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | ||
<foreignObject x="200" y="780" width="1100" height="300" > | <foreignObject x="200" y="780" width="1100" height="300" > | ||
− | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size:52px;font-weight:bold;color:black;">Illumination with violet light (400nm) re-activates | + | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size:52px;font-weight:bold;color:black;">Illumination with violet light (400nm) re-activates Dronpa fluorescence and also leads to the closed conformation of the construct.</p> |
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<rect x="70" y="70" rx="20" ry="20" width="900" height="270" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | <rect x="70" y="70" rx="20" ry="20" width="900" height="270" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | ||
<foreignObject x="70" y="70" width="900" height="270" > | <foreignObject x="70" y="70" width="900" height="270" > | ||
− | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size: | + | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size:47px;font-weight:bold;color:black;">Illumination with blue light (500nm) deactivates Dronpa fluorescence and thereby also multimerisation of the Dronpa domains.</p> |
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<rect x="950" y="700" rx="20" ry="20" width="950" height="340" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | <rect x="950" y="700" rx="20" ry="20" width="950" height="340" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | ||
<foreignObject x="950" y="700" width="950" height="340" > | <foreignObject x="950" y="700" width="950" height="340" > | ||
− | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size:52px;font-weight:bold;color:black;">In the induced open conformation of the construct, the | + | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size:52px;font-weight:bold;color:black;">In the induced open conformation of the construct, the Dronpa domains are only losely connected to the Cre, which thereby becomes active.</p> |
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<rect x="30" y="400" rx="20" ry="20" width="870" height="300" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | <rect x="30" y="400" rx="20" ry="20" width="870" height="300" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | ||
<foreignObject x="30" y="400" width="870" height="300" > | <foreignObject x="30" y="400" width="870" height="300" > | ||
− | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size: | + | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size:46px;font-weight:bold;color:black;">The RFP-luciferase reporter switch leads to expression of RFP if the memory system has not been activated by CREllumination.</p> |
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<rect x="1000" y="30" rx="20" ry="20" width="900" height="450" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | <rect x="1000" y="30" rx="20" ry="20" width="900" height="450" style="fill:#b3b3b3;stroke:black;stroke-width:5;opacity:0.78" /> | ||
<foreignObject x="1000" y="30" width="900" height="450" > | <foreignObject x="1000" y="30" width="900" height="450" > | ||
− | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size:52px;font-weight:bold;color:black;">After | + | <p xmlns="http://www.w3.org/1999/xhtml" style="font-size:52px;font-weight:bold;color:black;">After activation of the Cre protein it removes the loxp-RFP-loxp part of the reporter switch. This leads to expression of luciferase, which serves as the final reporter of the system.</p> |
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<g class="hover_group" opacity="0"> | <g class="hover_group" opacity="0"> | ||
− | <a xlink:href=" | + | <a xlink:href="https://2015.igem.org/Team:Tuebingen/Design?slide=3"> |
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− | <a xlink:href=" | + | <a xlink:href="https://2015.igem.org/Team:Tuebingen/Design?slide=1"> |
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− | <a xlink:href=" | + | <a xlink:href="https://2015.igem.org/Team:Tuebingen/Design?slide=1"> |
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− | <a xlink:href=" | + | <a xlink:href="https://2015.igem.org/Team:Tuebingen/Design?slide=2"> |
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− | To achieve the construction of a reversibly activatable Cre recombinase we want to apply the caging mechanism described by Zhou et al [ | + | To achieve the construction of a reversibly activatable Cre recombinase we want to apply the caging mechanism described by Zhou et al <a href="https://2015.igem.org/Team:Tuebingen/References">[Zhou 2012]</a>. This caging is performed by fusing a copy of a variant of the fluorescent protein Dronpa to both the C- and N-terminus of the Cre recombinase. Since this Dronpa variant is able to form monomers or dimers depending on illumination with light of different wavelengths, we hope that the dimerized form inhibits the activity of the Cre recombinase. </p> |
<p> | <p> | ||
Because our system only needs the expression of the caged Cre construct to be dependent on a sensor, it can be combined with almost all Biosensors that include a means of transcriptional control. This gives the system a wide variety of possible applications, especially in the context of the work of other iGEM teams.</p> | Because our system only needs the expression of the caged Cre construct to be dependent on a sensor, it can be combined with almost all Biosensors that include a means of transcriptional control. This gives the system a wide variety of possible applications, especially in the context of the work of other iGEM teams.</p> | ||
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</div> | </div> | ||
Latest revision as of 14:57, 9 November 2015
Project Description
A Biosensor Memory Module: Cre Sensor
To achieve the construction of a reversibly activatable Cre recombinase we want to apply the caging mechanism described by Zhou et al [Zhou 2012]. This caging is performed by fusing a copy of a variant of the fluorescent protein Dronpa to both the C- and N-terminus of the Cre recombinase. Since this Dronpa variant is able to form monomers or dimers depending on illumination with light of different wavelengths, we hope that the dimerized form inhibits the activity of the Cre recombinase.
Because our system only needs the expression of the caged Cre construct to be dependent on a sensor, it can be combined with almost all Biosensors that include a means of transcriptional control. This gives the system a wide variety of possible applications, especially in the context of the work of other iGEM teams.