Griffitts:Nodule Recovery

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Procedure

  • Place plant in a puddle of water
    • The lid of a Petri dish works nicely
  • Cut off the nodule using a sterile razor blade
  • Place nodule in a microcentrifuge tube with 500 μL dH2O
    • Be sure to label your tubes
  • Decant dH2O
  • Add 500 μL 75% ethanol
  • Wait 30 seconds
  • Decant ethanol
  • Add 500 μL 50% bleach
  • Wait 60 seconds
  • Decant bleach
  • Wash 2–3 times with sterile dH2O
  • Add 200 μL LB-10% glycerol
  • Macerate the nodule with a blue pestle
  • Pipette 5 μL to the appropriate selective medium and streak out with a single toothpick
    • This should be ~100 CFU
    • Be sure to label your plate
  • Freeze down the remaining solution in the –80°C freezer
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