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Team:Paris/Addressing overview2 strategy
From 2009.igem.org
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| - | Our strategy is to use clyA to export a protein to the outer-membrane of the cell. The protein fused to clyA will be incorporated into the vesicle during the vesiculation process. ClyA contain the signal peptide needed to be exported form the cytoplasm to the periplasm. The overall idea is to fused the protein of interest to clyA. | + | Our strategy is to use clyA to export a protein to the outer-membrane of the cell. The protein fused to clyA will be incorporated into the vesicle during the vesiculation process and it's also express on the surface. ClyA contain the signal peptide needed to be exported form the cytoplasm to the periplasm. The overall idea is to fused the protein of interest to clyA. |
So in a first time in order to see if our ClyA are in OMVs, we fused it we a RFP, for that we add a poly glycine linker to ClyA biobrick design for improving ClyA-RFP fusion. Moreover if we put RFP before ClyA, it seem that there is more fluorescence than ClyA before RFP under [article 1]. You can see the contruction here [[https://2009.igem.org/Team:Paris/Addressing_overview_Construction#Overview]] | So in a first time in order to see if our ClyA are in OMVs, we fused it we a RFP, for that we add a poly glycine linker to ClyA biobrick design for improving ClyA-RFP fusion. Moreover if we put RFP before ClyA, it seem that there is more fluorescence than ClyA before RFP under [article 1]. You can see the contruction here [[https://2009.igem.org/Team:Paris/Addressing_overview_Construction#Overview]] | ||
Revision as of 21:16, 20 October 2009
iGEM > Paris > ClyA > Our strategy
B. Our strategy
Our strategy is to use clyA to export a protein to the outer-membrane of the cell. The protein fused to clyA will be incorporated into the vesicle during the vesiculation process and it's also express on the surface. ClyA contain the signal peptide needed to be exported form the cytoplasm to the periplasm. The overall idea is to fused the protein of interest to clyA.
So in a first time in order to see if our ClyA are in OMVs, we fused it we a RFP, for that we add a poly glycine linker to ClyA biobrick design for improving ClyA-RFP fusion. Moreover if we put RFP before ClyA, it seem that there is more fluorescence than ClyA before RFP under [article 1]. You can see the contruction here [[1]]
Then if this test we could replace the RFP by the protein of interest for signal transduction,moreover this system couple to fec system can transduct a signal from outer membran to the cytoplasm.
pas toucher, j'essaye d'assembler ça et les foutre dans les bonnes partie merci^^---- Clya exprimé sur la membrane externe de la vésicule mais aussi dans, si exprimé a sur la surface il et transmembranaire?--> il a le c term a l intérieur et la partie n term a l ext d' ou la meilleur fluorescense, mais une meilleur conservation des peptide signal en c ter. On peut aussi penser qu on puisse adresser 2 sorte de signal, dans le periplasme si on greff notre protéine d' intérêt en c ter et sur les récepteurs de la membrane externe si on lui greffe la protéine en n ter.
