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Team:INSA-Lyon/Protocols/Transformation
From 2010.igem.org
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Revision as of 20:37, 25 August 2010
Protocols
Currently under construction
- Competent cells
- Transformation
- DNA extraction
- Digestion
- Ligation
- Measure of temperature and shaking speed influence
- Measure of osmotic pressure influence
- Granules extraction and intein cleavage
- Biofilms quantification
- Extra
Transformation
-
Always keep competent bacteria on ice before the heat shock in order to maintain competence.
- Add 1 µL of DNA (can be adjust with the DNA concentration, for example transform the entire ligation product) to 200 µl of competent cells. Mix carefully by moving the tip around.
- Incubate cells on ice for 30 min.
- Heat-shock cells for 2 min in a 42°C water bath.
- Incubate cells on ice for 5 min.
- Add 0.8 ml of room temperature LB.
- Incubate 1h at 37°C.
- Spread 100 µl of bacterial suspension on a LB agar plate containing the appropriate antibiotic and the rest in an other plate LB+AB
- Incubate overnight at 37°C
