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Team:DTU-Denmark/Notebook/4 July 2013
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===Preparation of SOB medium=== | ===Preparation of SOB medium=== | ||
Revision as of 13:19, 25 July 2013
Contents |
301
Main purpose
We want to view our cells throught a fluorescent microscope, in order to see if they have expressed GFP and RPF.
Hopefully we will see GFP in the periplasm and RFP inside the cell.
Setup an overnight culture of the 2 successful transformants so we can both MidiPrep and isolate periplasmic proteins tomorrow.
Procedure
Results
We managed to see expressed GFP and RFP
115
Main purpose
Preparation for making competent cells based on the protocol (link).
Who was in the lab
Henrike, Ariadni, Kashia, Natalia
Procedure
Preparation of SOB medium
Total volume 1 L
- 5 g yeast extract
- 20.05 g tryptone
- 0.58 g NaCl
- 0.19 g KCl
- 2.41 g MgSO4
Preparation of CCMB80 buffer (500 ml)
- KOAc :5 ml (0.98 gr)
- CaCl2.2H2O (5.9 g)
- MnCl2.4H2O (2 g)
- MgCl2.6H2O (1 g)
- 10% glycerol (50 ml)
Adjustment of pH (6.37) with 0.1 HCl
Preparation of LB agar
Conclusion from today
Cultivation of TOP10 cells in SOB
- 1 loop of TOP10 cells (from 14.06.2013)
- 10 ml of SOB medium
Incubate for 16 hours in 24 degrees and 37 degrees.
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