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Team:Paris/July 16
From 2008.igem.org
Results of transformations
- Positive controls from biobricks 2007, highest number of bacterias than the first try. It's probably due to the highest quantity of bacteria introduced (150µL instead 100µL).
- Transformations from biobricks 2008 don't succeed [[again]].
Preparation of competent cells
- Dilutions 1/200
- Culture during 3h --> DO= 0,15 / 4h --> DO= 0,13
- problem from the spectro, culture have been thrown.
Transformations Biobricks 2008
use of a new protocol --> directly competent cells
- Defroze competent cells on ice during 5'
- Mix 100µL of competent bacterias with 5µL of biobricks 08 (or 1µL for the positive control)
- Incubate 10' on ice
- Heat-shock the cells during 45-50" at 42°C (don't shake!!!)
- Put 2' on ice
- Add 900µL of cold SOC medium (4°C)
- Incubate 1h at 37°C under shaking (225rpm)
- Transfer in an eppendorf tube and spin at 5000rpm during 30"
- Remove 800µL of supernatant
- Resuspent the pellet in the 150-200µL left
- Spread on plates
- Incubate O/N at 37°C
