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Team:Heidelberg/Notebook/Sensing Group/Notebook/6thweek
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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)
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
Tuesdsay, 09/09/2008
- Miniprep of LuxQ colonies no. 1,2,3,5,6
- 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)
- Fusion-PCR for Fusion-1 and Fusion-2 constructs
