Team:Heidelberg/Notebook/Sensing Group/Notebook/6thweek

From 2008.igem.org

Revision as of 17:09, 26 October 2008 by Koljaschleich (Talk | contribs)

Back to the overview

Contents

Monday, 09/08/2008

  • Liquid Culture of 5 colonies from LuxQ Transformation and 1 from F1 transformation
  • Miniprep of Liquid Culture from previous Transformation (done by Chenchen). Incubated for two days @ RT and about 6 h @ 37 °C
  • Digestion of the Miniprep products with xbaI (Buffer2 + BSA, 1 ul enzyme @ 37 °C about 40min)
Digestion of LuxQ with xbaI yielded expected bands (4736 + 1204 bp) for all samples, except for no. 4
Sequencing @ GATC yielded correct sequence for LuxQ no. 1
  • PCR for LuxQ and LuxP with Phusion (25 µl Mastermix, 0.5 µl each Primer, 0(1) µl DMSO, 24(23) µl H20, 0.5 µl Template)
  • PCR Programme: 5 min @ 98 °C || 10 s @ 98 °C | 30 s @ 55 °C | 30 s @ 72 °C || 5 min @ 72 °C | 4 °C
PCR for LuxQ and LuxP. LuxQ PCR did not work, but first PCR for LuxP was positive.

Tuesdsay, 09/09/2008

  • Miniprep of LuxQ colonies no. 1,2,3,5,6
HD 080909-PCR Miniprep-LuxQ 2.png
HD 080909-PCR Miniprep-LuxQ 1.png


  • PCR for LuxQ fragment 1 and 2, Tar fragment 1 and 2 for Phusion receptor with Phusion-Polymerase
  • PCR Programme: 30 s @ 98 °C || 10 s @ 98 °C | 30 s @ 55 °C |30 s @ 72 °C || 5 min @ 72 °C | 4 °C (30 cycles)
PCR of LuxQ and Tar constructs for fusion receptor. LuxQ is negative
PCR of LuxQ constructs for fusion


  • Fusion-PCR for Fusion-1 and Fusion-2 constructs
Fusion-PCR
  • O/N Culture of 10 LuxP in pDK48 colonies

Wednesday, 09/10/2008

Thursday, 09/11/2008

Friday, 09/12/2008

Go to 7th week