Team:Rice University/Notebook/12 June 2008

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=Thursday 12 June=
=Thursday 12 June=
*Taylor Stevenson
*Taylor Stevenson
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**Transformation From New Registry (Paper Punches)-[[Team:Rice University/Electroporation|Electroporation]] and [[Team:Rice_University/Heat_Shock|Heat Shock]] of [http://partsregistry.org/Part:BBa_I744204|BBA_I744204] located on [http://partsregistry.org/Part:pSB1A2 pSB1A2] and [http://partsregistry.org/Part:BBa_I13521|BBA_I13521] located on [http://partsregistry.org/Part:pSB1A2 pSB1A2] into XL1-Blue MR electrocompetent cells.
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**Transformation From New Registry (Paper Punches)-[[Team:Rice University/Electroporation|Electroporation]] and [[Team:Rice_University/Heat_Shock|Heat Shock]] of [http://partsregistry.org/Part:BBa_I744204 BBa_I744204] located on [http://partsregistry.org/Part:pSB1A2 pSB1A2] (positive control) and [http://partsregistry.org/Part:BBa_I13521 BBa_I13521] located on [http://partsregistry.org/Part:pSB1A2 pSB1A2] into XL1-Blue MR electrocompetent and XL1-Blue chemically competent cells.
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*#Added 2uL of DNA solution obtained by soaking DNA punch in (10:1) Tris:EDTA buffer incubated @ 42*C for 20min.
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**#Added 1uL of stock miniprepped [http://partsregistry.org/Part:BBa_I744204 BBa_I744204] DNA solution and 2uL obtained by soaking [http://partsregistry.org/Part:BBa_I13521 BBa_I13521] DNA punch in (10:1) Tris:EDTA buffer incubated @ 50*C for 30min to both chemical and electrocompetent cells.
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*#Electropulse parameters: 1.8 kV & 5.4 ms
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**# Electropulse parameters for the miniprepped plasmid and extracted plasmid where 1.8kV & 5.2ms and 1.8kV & 5.0ms respectively.
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*#Plated 100uL and 20uL of transformed cells as well as 20uL of  1/100 and 1/10,000 dilutions in sterile 50% glycerol.  Plates were cultured O/N.  
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**# Plated 100uL of each the transformed and cultured cells on LB/Amp plates.  Plates were cultured O/N @ 37*C.
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**'''Result'''-no colonies were observed at any dilution
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***'''Result'''-no colonies were observed on plates from the extracted DNA while >>1,000 colonies grew on plates resulting from miniprepped DNA.
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**Obtaining a lambda lysogen
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***VCS257 cells were prepared for phage infection as specified in packaging manual [https://static.igem.org/mediawiki/2008/6/6a/Packaging_Extract.pdf] and infected with phage at roughly a 1/10 pfu/cfu ratio.  Resulting phage/bacteria mixture was incubated @ 37*C for 20 min and then streaked onto an LB plate.  Plate was incubated @ 30*C O/N. 
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***'''Result'''-approximately 100 possibly lysogenic colonies grew on plate.
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Latest revision as of 20:43, 16 July 2008

Thursday 12 June

  • Taylor Stevenson
    • Transformation From New Registry (Paper Punches)-Electroporation and Heat Shock of [http://partsregistry.org/Part:BBa_I744204 BBa_I744204] located on [http://partsregistry.org/Part:pSB1A2 pSB1A2] (positive control) and [http://partsregistry.org/Part:BBa_I13521 BBa_I13521] located on [http://partsregistry.org/Part:pSB1A2 pSB1A2] into XL1-Blue MR electrocompetent and XL1-Blue chemically competent cells.
      1. Added 1uL of stock miniprepped [http://partsregistry.org/Part:BBa_I744204 BBa_I744204] DNA solution and 2uL obtained by soaking [http://partsregistry.org/Part:BBa_I13521 BBa_I13521] DNA punch in (10:1) Tris:EDTA buffer incubated @ 50*C for 30min to both chemical and electrocompetent cells.
      2. Electropulse parameters for the miniprepped plasmid and extracted plasmid where 1.8kV & 5.2ms and 1.8kV & 5.0ms respectively.
      3. Plated 100uL of each the transformed and cultured cells on LB/Amp plates. Plates were cultured O/N @ 37*C.
      • Result-no colonies were observed on plates from the extracted DNA while >>1,000 colonies grew on plates resulting from miniprepped DNA.
    • Obtaining a lambda lysogen
      • VCS257 cells were prepared for phage infection as specified in packaging manual [1] and infected with phage at roughly a 1/10 pfu/cfu ratio. Resulting phage/bacteria mixture was incubated @ 37*C for 20 min and then streaked onto an LB plate. Plate was incubated @ 30*C O/N.
      • Result-approximately 100 possibly lysogenic colonies grew on plate.






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