Griffitts:Nodule Recovery

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