Team:Unimelb/Experiments

Experiments

SOC MEDIUM
  • To 950 mL of deionised water, add:
    Tryptone 20g
    Yeast Extract 5g
    NaCl 0.5g
  • Dissolve the solutes by placing the flask on the shaker
  • Add 10 mL of a 250 mM solution of KCl
  • Add 5 mL of sterile solution of 2 M MgCl2 once solution is 60°C or less
  • Fill to 980 mL with deionised water
  • Autoclave the SOC medium at 121°C for 20 minutes for sterilisation
  • Add 20 mL of sterile 1 M solution of glucose
  • Sterilise by passing it through a 0.22-µm filter
  • Store the SOC medium at room temperature and for long term storage, store at 4°C.

LB MEDIUM (1L LIQUID)
  • For 1L LB medium, the correct amounts are:
    NaCl 10g
    Peptone (or tryptone) 10g
    Yeast extract 5g
  • Collect them in in a bottle and add 1L of dH2O
  • Autoclave the LB medium at 121°C for 20 minutes for sterilisation.

LB AGAR
  • For 200 mL LB-agar, the correct amounts are:
    NaCl 2g
    Peptone (or tryptone) 2g
    Yeast extract 1g
    Bacto-agar 3g
  • Collect them in a bottle and add 200 mL of H2O
  • Autoclave the LB medium at 121°C for 20 minutes for sterilisation.

Note: With 200 mL LB-agar 8 plates can be made.


AMPICILLIN STOCK (100 mg/ml)
  • Dissolve 0.5 g ampicillin in 5 mL Milli Q.
  • Mix/vortex so that all the ampicillin goes into solution.
  • Filter into a falcon tube using a syringe and a 0.22 μm filter for sterilization.
  • Aliquot into smaller Eppendorf tubes.
  • Store at -20°C under dark conditions.

CHLORAMPHENICOL STOCK (25mg/ml)
  • Dissolve 0.125 g chloramphenicol in 5 mL Milli Q.
  • Mix/vortex so that all the ampicillin goes into solution.
  • Filter into a falcon tube using a syringe and a 0.22 μm filter for sterilization.
  • Aliquot into smaller Eppendorf tubes.
  • Store at -20°C under dark conditions.

CELL COMPETENCY TEST

For cell competency test we used iGEM Competent Cell Test Kit according to the instructions provided by http://parts.igem.org/Help:2018_Competent_Cell_Test_Kit .