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Revision as of 00:05, 5 October 2013
Protein Expression
Materials
Equipment
- - Incubation shaker
- - Photometer
Chemicals & consumables
- - E. coli BL21 DE3
- - dYT Medium
- - IPTG
- - 100 ml and 3 l flasks
- - ice
Procedure
- Inoculation of 50 mL DYT medium in the 100 mL flask with E. coli BL21 DE3 containing pPR-IBA2
plasmid with the sequence for the respective part to be expressed.
- Incubation at 180 repulsion per minute (rpm) at 30°C to an OD600= 4
- Transferation of the starter culture into 1 L DYT medium in a 3 L flask resulting in an OD600= 0.2
- Incubation to an OD600= 0.6 at 180 rpm and 30°C.
- Incubation for 15 minutes on ice.
- Induction of the proteinexpression with 20 mL of IPTG (stock conentration 1M).
- Incubation of the cell suspension over night at 180 rpm at 30°C.
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