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