Team:EPF Lausanne/Notebook/week1

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= Week 1 =
 +
 +
=12-07-2010=
 +
* OD measurement of Asaia’s culture
 +
* Chemical competence for E.Coli DB3.1 (step : Day 3)
 +
* PCR (Asaia ORI + primers)=12-07-2010=
 +
* OD measurement of Asaia’s culture
 +
* Chemical competence for E.Coli DB3.1 (step : Day 3)
 +
* PCR (Asaia ORI + primers)
 +
* Made GLY agar plates without antibiotics -> failed (medium was too old)
 +
* Asaia O/N culture without antibiotics in order to recover some WT
 +
* Text for the sponsor
 +
 +
 +
=13-07-2010=
 +
* Chemical competence for E.Coli DB3.1 (step : Day 4) -> stored at -80°C
 +
* Preparation of GLY Medium / LB Medium / SOC Medium / TAE 1X / TAE 0.5X
 +
* Run a gel for the PCR -> failed (mix of two different kits)
 +
* Preparation of plates (GLY Agar / LB+Kan Agar / LB+Amp Agar)
 +
* Learned to autoclave
 +
* Restarted PCR (Asaia ORI + primers)
 +
* Plated Asaia (O/N culture)
 +
* Choice of Antibiotics Biobricks resistance (Kan / Amp / Tet / Chl) from the iGem Kit and preparation of them (streak out)
 +
* Transformation of E.coli DB3.1 with pUC19 to check its competence
 +
* Transformation of E.coli DH5 with p1003 (Kan) / p1002 (Amp) / p1004 (Chl) / p1005 (Tet)
 +
* E.coli DB3.1 and DH5 were plated on LB+Amp Agar plates (Kan was plated on LB+Kan Agar plate)
 +
* Ordered material
 +
* Wiki brainstorming
 +
* Protocols printing and organization
 +
 +
 +
=14-07-2010=
 +
* Run a gel for the PCR of the previous day -> worked
 +
* Purification of PCR’s product -> ok
 +
* Preparation of the BBa_151020 (streak out)
 +
* Competence Calculation of E.Coli DB3.1 (+pUC10, Amp) -> ok
 +
* Look at Asaia with microscope -> we have cells (pictures)
 +
* Growth Curve for Asaia (OD measurement each hour) -> both manually (ok) + with the plate reader
 +
* Plated Asaia to recover some WT (streak out)
 +
* Preparation of Asaia Culture (for Lemaître experiment) with Kan
 +
* Protocol LateX template
 +
* O/N culture of the E.coli DH5 transformed with the BB Antibiotics resistance (4x) -> miniprep
 +
* Learning HTML for the wiki
 +
* Text for the project presentation due to iGem
 +
* Writing Protocols of basic procedures for the wiki page
 +
* Culture of Asaia + Kan
 +
 +
 +
=15-07-2010=
 +
* MaxiPrep of Lemaître’s culture
 +
* Purification of the BB resistances from the E.Coli cultures
 +
* Protocol for cloning the Asaia Vector
 +
* Enzyme digestion of the BB resistances and Asaia_ORI
 +
* Cloning (Asaia_ORI + Vector/ Asaia_ORI + BB + Vector)
 +
* Wiki text
 +
* Preparation of LB agar + Cm
 +
* Electrophoresis of resistance fragments (Works!!!) and extraction
 +
* Purification of extracted DNA from agarose gel
 +
 +
 +
=16-07-2010=
 +
* ligation Asaia origin BB + vector
 +
* ligation Asaia origin BB + Kan + vector
 +
* ligation Asaia origin BB + Amp + vector
 +
* ligation Asaia origin BB + Cm + vector
 +
* ligation Asaia origin BB + Tet + vector
 +
* Extraction of the OD curve data (values)
 +
* Replating Asaia on plates with and without kanamicin in order to recover the WT Asaia (only WT do not grow on kanamicin)
 +
* Feeding and infection of Drosophila (WT and immunodeficient redish mutants) with bacteria Ecc15 and Asaia in Prof. Lemaitre Lab
 +
* Fluorescence microscopy -> presence of fluorescence in the body/gut as expected
 +
* Transformation of E-Coli with the ligation products and plating the transformed bacteria
 +
 +
 +
 +
 +
{{EPFL_2010_bottom_wrap}}
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Latest revision as of 19:47, 24 October 2010









Contents

Week 1

12-07-2010

  • OD measurement of Asaia’s culture
  • Chemical competence for E.Coli DB3.1 (step : Day 3)
  • PCR (Asaia ORI + primers)=12-07-2010=
  • OD measurement of Asaia’s culture
  • Chemical competence for E.Coli DB3.1 (step : Day 3)
  • PCR (Asaia ORI + primers)
  • Made GLY agar plates without antibiotics -> failed (medium was too old)
  • Asaia O/N culture without antibiotics in order to recover some WT
  • Text for the sponsor


13-07-2010

  • Chemical competence for E.Coli DB3.1 (step : Day 4) -> stored at -80°C
  • Preparation of GLY Medium / LB Medium / SOC Medium / TAE 1X / TAE 0.5X
  • Run a gel for the PCR -> failed (mix of two different kits)
  • Preparation of plates (GLY Agar / LB+Kan Agar / LB+Amp Agar)
  • Learned to autoclave
  • Restarted PCR (Asaia ORI + primers)
  • Plated Asaia (O/N culture)
  • Choice of Antibiotics Biobricks resistance (Kan / Amp / Tet / Chl) from the iGem Kit and preparation of them (streak out)
  • Transformation of E.coli DB3.1 with pUC19 to check its competence
  • Transformation of E.coli DH5 with p1003 (Kan) / p1002 (Amp) / p1004 (Chl) / p1005 (Tet)
  • E.coli DB3.1 and DH5 were plated on LB+Amp Agar plates (Kan was plated on LB+Kan Agar plate)
  • Ordered material
  • Wiki brainstorming
  • Protocols printing and organization


14-07-2010

  • Run a gel for the PCR of the previous day -> worked
  • Purification of PCR’s product -> ok
  • Preparation of the BBa_151020 (streak out)
  • Competence Calculation of E.Coli DB3.1 (+pUC10, Amp) -> ok
  • Look at Asaia with microscope -> we have cells (pictures)
  • Growth Curve for Asaia (OD measurement each hour) -> both manually (ok) + with the plate reader
  • Plated Asaia to recover some WT (streak out)
  • Preparation of Asaia Culture (for Lemaître experiment) with Kan
  • Protocol LateX template
  • O/N culture of the E.coli DH5 transformed with the BB Antibiotics resistance (4x) -> miniprep
  • Learning HTML for the wiki
  • Text for the project presentation due to iGem
  • Writing Protocols of basic procedures for the wiki page
  • Culture of Asaia + Kan


15-07-2010

  • MaxiPrep of Lemaître’s culture
  • Purification of the BB resistances from the E.Coli cultures
  • Protocol for cloning the Asaia Vector
  • Enzyme digestion of the BB resistances and Asaia_ORI
  • Cloning (Asaia_ORI + Vector/ Asaia_ORI + BB + Vector)
  • Wiki text
  • Preparation of LB agar + Cm
  • Electrophoresis of resistance fragments (Works!!!) and extraction
  • Purification of extracted DNA from agarose gel


16-07-2010

  • ligation Asaia origin BB + vector
  • ligation Asaia origin BB + Kan + vector
  • ligation Asaia origin BB + Amp + vector
  • ligation Asaia origin BB + Cm + vector
  • ligation Asaia origin BB + Tet + vector
  • Extraction of the OD curve data (values)
  • Replating Asaia on plates with and without kanamicin in order to recover the WT Asaia (only WT do not grow on kanamicin)
  • Feeding and infection of Drosophila (WT and immunodeficient redish mutants) with bacteria Ecc15 and Asaia in Prof. Lemaitre Lab
  • Fluorescence microscopy -> presence of fluorescence in the body/gut as expected
  • Transformation of E-Coli with the ligation products and plating the transformed bacteria



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