Difference between revisions of "Haynes Lab:Notebook/ASU iGEM"

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===6-13-12===
 
* Transformation (LSE)
 
:* Transformed DNA:
 
::* lacZ (well 4:12G, I732019)
 
::* p + lacO (well 1:6G, R0011)
 
:* Cells: DH5 alpha (donated)
 
:* Protocol from: http://openwetware.org/wiki/Haynes:Assembly101 (30 minute transformation)
 
:* Controls: puc19, no DNA (8 plates)
 
* Transformation (istb4, Abhi)
 
:* Transformed DNA:
 
:* Cells:
 
:* Protocol from:
 
:* Controls:
 
*DH5a Chemically Competent cell prep
 
:*Grew 2 seed colonies from streak plate in LB no amp
 
:*Grew controls to test for contamination
 
::*Both Seed colonies grew, no contamination present
 
  
 
===6-14-12===
 
===6-14-12===

Revision as of 17:09, 3 July 2012

<!-- sibboleth --><div id="lncal1" style="border:0px;"><div style="display:none;" id="id">lncal1</div><div style="display:none;" id="dtext">06/07/2012,06/08/2012,06/12/2012,06/13/2012,06/14/2012,06/15/2012,06/16/2012,06/17/2012,06/19/2012,06/20/2012,06/21/2012,06/22/2012,06/26/2012,06/27/2012,06/28/2012,07/02/2012,07/03/2012,07/24/2012,07/25/2012,07/26/2012,07/27/2012,07/28/2012,07/30/2012,07/31/2012,08/03/2012,08/10/2012,08/13/2012,08/14/2012,08/15/2012,08/16/2012</div><div style="display:none;" id="page">Haynes Lab:Notebook/ASU iGEM</div><div style="display:none;" id="fmt">yyyy/MM/dd</div><div style="display:none;" id="css">OWWNB</div><div style="display:none;" id="month"></div><div style="display:none;" id="year"></div><div style="display:none;" id="readonly">Y</div></div>

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Project Description/Abstract

Team Chimeric Reporter:

  • Hyder Hussain
  • Abhinav Markus
  • Ryan Muller
  • Ethan Ward

Team Cell Surface Reporter:

  • Nisarg Patel
  • Ryan Muller
  • Rohit Rajan
  • Ellen Qin
  • Melinda Jenner
  • Joe Barth

BioBricks

Magainin Reporter

  • Magainin
  • Magainin + 2xGS Linker + His-Tag
  • Bgal fragment (a-1)
  • Bgal fragment (a-4)
  • Bgal fragment (w)
  • Bgal fragment (1-w)
  • Magainin + 2aa GS Linker + His-Tag + 10aa GS Linker + Bgal fragment (a-1)
  • Magainin + 2aa GS Linker + His-Tag + 10aa GS Linker + Bgal fragment (a-4)
  • Magainin + 2aa GS Linker + His-Tag + 10aa GS Linker + Bgal fragment (w)
  • Magainin + 2aa GS Linker + His-Tag + 10aa GS Linker + Bgal fragment (1-w)
  • His-Tag + 10aa GS Linker + Bgal fragment (a-1)
  • His-Tag + 10aa GS Linker + Bgal fragment (a-4)
  • 13. His-Tag + 10aa GS Linker + Bgal fragment (w)
  • 14. His-Tag + 10aa GS Linker + Bgal fragment (1-w)

Notes

6-14-12

  • Competent cell prep
  • Prepared CaCl2 buffer solution and CaCl2 glycerol buffer solution
  • Grew seed colony in 400mL LB no amp

6-15-12

  • Competent cell prep
  • Centrifuged falcon test tubes containing liquid colonies
  • Resuspended in CaCl2 buffer solution and incubated for 15 mins
  • Centrifuged and resuspended in CaCl2 glycerol buffer solution
  • Chilled overnight

6-16-12

  • Competent cell prep
  • Aliquotted 200uL into test tubes
  • Stored in -80C

6-17-12

  • Streak plated prepared competent cells on LB no amp plate
  • Colonies observed

6-19-12

  • Transformation (LSE)
  • Transformed DNA:
  • T7 promoter BBa_I712074
  • Constitutive promoter BBa_J23102
  • Made 50 LB Amp plates.

6-20-12

  • Plated negative control on LB Amp plate
  • Liquid cultures of T7 promoter and constitutive promoter
  • Transformation (LSE)
  • Transformed DNA:
  • RBS (well 1:1H BBa_B0030)
  • TetR GFP (well 2:8A Part:BBa_I13522)

6-21-12

  • Made Liquid Cultures of E.coli transformed with RBS B0030
  • Made Liquid Cultures of E.coli transformed with TetR GFP
  • miniprepped and nanodropped T7 promoter BBa_I712074 and Constitutive promoter BBa_J23102 liquid cultures
  • liquid cultures:
  • RBS1
  • RBS2 (duplicate_
  • GFP1
  • puc19
  • negative controls
  • 5 ml LB amp
  • overnight cultures
  • replated GFP1 & 2 (duplicates)
  • Nanodropped plasmid DNA samples
  • Constitutive promoter 1: __ng/uL
  • Constitutive promoter 2: __ng/uL
  • T7 promoter 1: __ng/uL
  • T7 promoter 2: __ng/uL

6-22-12

  • Miniprepped liquid cultures: RBS (well 1:1H BBa_B0030) and TetR GFP (well 2:8A Part:BBa_I13522)

6-26-12

  • Transformation:
  • Transformed DNA:
  • double terminator (B0017, 2:6K)
  • T7 RNA polymerase (I715038, 2:15C)
  • puc19, negative control

6-27-12

  • 6-26 transformation results:
  • Controls correct
  • 2x terminator: ~19 colonies
  • RNA pol: 1 colony
  • Liquid cultures including controls

6-28-12

  • Miniprepped double terminator (B0017, 2:6K) and T7 RNA polymerase (I715038, 2:15C) liquid cultures

7-2-12

  • Cleaned up liquid waste
  • Made SOB media
  • Finalized oligos for magainin construct

7-3-12

  • Autoclaved SOB media
  • Added glucose to make SOC media
  • Nanodropped double terminator (B0017, 2:6K) [DT1: 24.5, DT2: 29.6] and T7 RNA polymerase (I715038, 2:15C) [P1: 64.6, P2: 55.3] liquid cultures