Team:INSA-Lyon/Protocols/Transformation

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Revision as of 13:20, 10 September 2010







Protocols


Currently under construction



  1. Competent cells
  2. Transformation
  3. DNA extraction
  4. Digestion
  5. Ligation
  6. Measure of temperature and shaking speed influence
  7. Measure of osmotic pressure influence
  8. Granules extraction and intein cleavage
  9. Biofilms quantification
  10. 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

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