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| {{EPF-Lausanne09}} | | {{EPF-Lausanne09}} |
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| <div CLASS="epfltrick">__TOC__ | | <div CLASS="epfltrick">__TOC__ |
| </div><div CLASS="epfl09"> | | </div><div CLASS="epfl09"> |
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- | == The BphP / PpsR system ==
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- | PpsR1 is a redox sensitive activator. It binds DNA under anaerobic conditions, and forms a tetramer via a disulfide bond. This interaction is ablated in the mutant PpsR1-C429S; meaning that we should be able to mimic an anaerobic system even under aerobic conditions.
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| + | <font size="6" color="#007CBC"><i>R-BphP project </i></font> |
| + | </center></html> |
| + | <br> |
| + | ---- |
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| + | <br> |
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- | PpsR2 is a transcriptional repressor, regulated by BphP.
| + | This part of the project was cancelled, due to lack of time. You can read what our idea was on [https://2009.igem.org/Team:EPF-Lausanne/Brainstorming the brainstorming session] |
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- | PpsR1 and PpsR2 bind to TGTN_12ACA possibly arranged in tandem with a 7 base spacer. The affinity for both PpsRs are around 100nM.
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- | BphP (or BrBphP for the Bradyrhizobium variant) is sensitive to far-red light (~770nm) and controls PpsR1.
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- | For more info, see [http://www.jbc.org/cgi/content/full/279/43/44407].
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- | Similar constituents could possibly be derived from R. Palustris as well with the following model derived from [http://www3.interscience.wiley.com/cgi-bin/fulltext/118512333/HTMLSTART]:
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- | [[Image:R._palustris_circuit.gif]]
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- | ==To find==
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- | - Define genetic circuit <big>'''Tú'''</big>
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- | - Look for exact wavelength for the corresponding light receptor<big><span style="color:Blue"> Basile</span></big>
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- | - Sequences of the genes involved in the pathway (minimal genes to => [http://www.ncbi.nlm.nih.gov/ Pubmed])
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- | [[Media: ORS278_BrBphP_genomic_seq.txt|ORS278 BrBphP genomic sequence]]
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- | [[Media: ORS278_PpsR1_genomic_seq.txt|ORS278 PpsR1 genomic sequence]]
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- | [[Media: ORS278_PpsR2_genomic_seq.txt|ORS278 PpsR2 genomic sequence]]
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- | [[Media: ORS278_PpsR1-C429S_protein_seq.txt|ORS278 PpsR2-C429S protein sequence]]
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- | [[Media: hmuO_ORS278 Brad.txt|ORS278 hmuO genomic sequence]]
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- | [[Media: ppsR1_prot_RPalustris.txt| R.Palustris CGA009 ppsR1 protein sequence]]
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- | [[Media: ppsR1_RPalustris.txt| R.Palustris CGA009 ppsR1 genomic sequence]]
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- | [[Media: ppsR2_prot_RPalustris.txt| R.Palustris CGA009 ppsR2 protein sequence]]
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- | [[Media: ppsR2_RPalustris.txt| R.Palustris CGA009 ppsR2 genomic sequence]]
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- | [[Media: CAE001 bphP1.txt| R.Palustris CAE001 bphP1 genomic sequence]]
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- | [[Media: CGA009 hmuO.txt| R.Palustris CGA009 hmuO genomic sequence]]
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- | [[Image: bphp_organisation.gif|500px|thumb|center| Organization of the genes downstream the puf operon in Bradyrhizobium and Rps. palustris. BphP: bacteriophytochrome gene; hmuO: heme oxygenase gene.]]
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- | ==Implementation==
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- | There are two modes in which we can use the system. The minimal complement requires PpsR2 and its regulator BphP. We can generate a hybrid system that uses a well known activator or transcription or a constitutively active promoter which will be repressed via PpsR2 upon exposure to far-red light (770nm).
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- | It is also possible to reconstitute the entire system including PpsR1-C429S, which will serve as the activator and will be de-coupled from the oxidative state of the cell due to the ablation of the disulfide bond formation.
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- | ==To do==
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- | * contact Eric Giraud (giraud@mpl.ird.fr) and ask if he could send us the following material:
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- | ** Bradyrhizobium ORS278
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- | ** The following plasmids:
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- | *** pBAD::ppsR1
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- | *** pBAD::ppsR2
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- | *** pGEM-T::ppsR2/BrbphP
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- | ** the PpsR1-C429S construct
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- | ==Found==
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- | - Bacterium: Bradyrhizobium can be ordered on [http://www.lgcstandards-atcc.org/LGCAdvancedCatalogueSearch/ProductDescription/tabid/1068/Default.aspx?ATCCNum=10244&Template=bacteria ATCC]
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- | ----
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- | <!--
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- | {| style="color:#1b2c8a;background-color:#0c6;" cellpadding="3" cellspacing="1" border="1" bordercolor="#fff" width="62%" align="center"
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- | !align="center"|[[Team:EPF-Lausanne|Home]]
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- | !align="center"|[[Team:EPF-Lausanne/Team|The Team]]
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- | !align="center"|[[Team:EPF-Lausanne/Project|The Project]]
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- | !align="center"|[[Team:EPF-Lausanne/Parts|Parts Submitted to the Registry]]
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- | !align="center"|[[Team:EPF-Lausanne/Modeling|Modeling]]
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- | !align="center"|[[Team:EPF-Lausanne/Notebook|Notebook]]
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- | !align="center"|[[Team:EPF-Lausanne/Lectures|Lectures]]
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- | !align="center"|[[Team:EPF-Lausanne/Team Management|Team Management]]
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- | !align="center"|[[Team:EPF-Lausanne/References|References]]
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- | |}-->
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| </div><div CLASS="epfl09bouchon"></div> | | </div><div CLASS="epfl09bouchon"></div> |