Team:Heidelberg/Notebook/Sensing Group/Notebook/8thweek

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   Home    Team   Overview    Advisors  </a></li>  Undergraduates  </a></li>  University  </a></li>  DKFZ  </a></li>  BioQuant  </a></li>  BioRegion Rhein-Neckar  </a></li> </ul> </li>  Project</a> <ul class="DropDownMenu" id="MB1-DDM1">  Overall Project  </a></li>  Material & Methods  </a> <li><a href="http://2008.igem.org/Team:Heidelberg/Project/Sensing"> Sensing  </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Project/Killing_I"> Killing I  - Phages  </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Project/Killing_II"> Killing II - Colicin  </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Project/Visualization"> Visualization  </a></li> </ul> </li>

<li> <a href="http://2008.igem.org/Team:Heidelberg/Parts" style="color: white">Parts</a> <ul class="DropDownMenu" id="MB1-DDM2"> <li><a href="http://2008.igem.org/Team:Heidelberg/Parts"> Submitted Parts  </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Parts/Characterization"> Characterization  </a></li> </ul> </li> <li> <a href="http://2008.igem.org/Team:Heidelberg/Modeling" style="color: white">Modeling</a> <ul class="DropDownMenu" id="MB1-DDM3"> <li><a href="http://2008.igem.org/Team:Heidelberg/Modeling"> Overview  </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Modeling/Chemotaxis"> Chemotaxis-Colicin  </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Modeling/Phage"> Phage Dynamics model  </a></li> </ul> </li> <li> <a href="http://2008.igem.org/Team:Heidelberg/Notebook/Overview" style="color: white">Notebook</a> <ul class="DropDownMenu" id="MB1-DDM5"> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/Sensing_Group/Notebook"> Sensing >  </a> <ul class="SideMenu" id="MB1-DDM2-SM1"> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/Sensing_Group/Cloning"> Cloning strategy </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/Sensing_Group/Notebook"> Notebook </a></li> </ul> </li> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/Killing_I/Notebook"> Killing I - Phages >  </a> <ul class="SideMenu" id="MB1-DDM2-SM2"> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/Killing_I/Cloning"> Cloning strategy </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/Killing_I/Notebook"> Notebook </a></li> </ul> </li> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/Killing_II/Notebook"> Killing II - Colicin >  </a> <ul class="SideMenu" id="MB1-DDM2-SM3"> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/Killing_II/Cloning"> Cloning strategy </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/Killing_II/Notebook"> Notebook </a></li> </ul> </li> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/visualization"> Visualization  </a> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/material"> Material & Methods  </a> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/team_meetings"> Team Meetings  </a> <li><a href="http://2008.igem.org/Team:Heidelberg/Notebook/seminar"> Seminar on Synthetic Biology  </a> </ul> </li> <li style="width: 160px"> <a href="http://2008.igem.org/Team:Heidelberg/Human_Practice/Project_Overview" style="color: white">Human Practice</a> <ul class="DropDownMenu" id="MB1-DDM4"> <li><a href="http://2008.igem.org/Team:Heidelberg/Human_Practice/Project_Overview"> Project Overview  </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Human_Practice/Phips_the_Phage"> Phips the Phage  </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Human_Practice/Essay"> Essay  </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Human_Practice/Surveys"> Surveys  </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Human_Practice/Open_Day"> Open Day  </a></li> <li><a href="http://2008.igem.org/Team:Heidelberg/Human_Practice/Nobel_Prize"> Nobel Prize  </a></li> </ul> </li>

<li> <a href="http://2008.igem.org/Team:Heidelberg/Sponsors" style="color: white">Sponsors</a> </li> </ul>

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Back to the overview

Monday, 09/22/2008

 * test digestion of F1_LuxP_NdeI (F1 cloned in pDK48 with NdeI), F2_LuxP_NdeI, F1_LuxP_KpnI (F1 cloned in pDK48 with KpnI) and F2_LuxP_KpnI (F2 cloned in pDK48 with KpnI) with NcoI (NEBuffer 3)
 * Transformation of Fusion constructs (F1(NdeI)_LuxP_pDK48, F1(KpnI)_LuxP_pDK48 and F2(NdeI)_LuxP_pDK48) into MG1655 and HCB33 cells
 * Preparation of cultures for Maxipreps of Fusion constructs

Tuesday, 09/23/2008

 * Maxiprep of LuxP-Fusion constructs (F1(NdeI)_LuxP_pDK48, F1(KpnI)_LuxP_pDK48 and F2(NdeI)_LuxP_pDK48)
 * Cloning of Fusion_LuxP in pBAD33 with BamHI/PstI
 * digestions of Fusion constructs and pBAD33 as described in gel image descriptions

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 * O/N culture of Fusion-LuxP-pDK48 in HCB33 and MG1655 in TB-Kan
 * Transformation of Fusion-LuxP-pBAD33 constructs into DH5a

Wednesday, 09/24/2008
Correct sequence for both Fusion constructs confirmed
 * Glycerol-Stock of Fusion1-LuxP-pDK48 and Fusion2-LuxP-pDK48
 * O/N culture of Fusion-LuxP-pDK48 in HCB33 and MG1655 in LB
 * O/N culture of LuxS and GFP Plasmid from Marcus
 * Re-Sequencing of Fusion-1 and Fusion-2

Thursday, 09/25/2008

 * Miniprep of Fusion-LuxP-pDK48 constructs, LuxS, GFP Plasmid from Marcus
 * Competent Cells (Top-10, UU1250)
 * Picked colonies of Fusion-LuxP-pBAD33

Friday, 09/26/2008

 * Miniprep of Fusion-LuxP-pBAD33 constructs and Digestion with HindIII (NEBuffer2)
 * 5 µl DNA
 * 1µl NEBuffer 2
 * 0.5µl HindIII
 * 3.5µl H2O
 * Expected Fragments
 * Fusion1-LuxP-pBAD33: 7616bp and 769bp
 * Fusion2-LuxP-pBAD33: 7631bp and 769bp
 * pBAD33 only: 5360bp
 * Double Transformations of Fusion-LuxP-pBAD33 and pDK4 into UU1250 (2 Amp-CM plates) as well as LuxS and pDK6 into DH5a (Amp-Kan plate(s))
 * Transformation of pDK4 into DH5a (CM plate)
 * Preparation of 2x 3mL LB-CM-Amp, 2x 5mL TB-CM-Amp, 1x 3mL LB-Amp-Kan, 1x 5mL TB-Amp-Kan, 1x 5mL LB-CM

Sunday, 09/28/2008

 * Picked colony of each Fusion-LuxP-pBad/pDK4 UU1250 cells and inoculated in 5 mL TB and 5 mL LB medium
 * No colonies on LuxS/pDK6 DH5a cell plate. 5 mL liquid culture of LuxS DH5a cells.
 * No colonies on pDK4 cell plate. Waiting until tomorrow

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