Difference between revisions of "Mimulus:Seed Sterilization"

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(Supplies)
(Methods)
 
Line 12: Line 12:
 
#Immediately add 600 uL of Water. Do only 2 tubes at a time, because the alcohol treatment is harsh and might affect germination if is left on seeds too long
 
#Immediately add 600 uL of Water. Do only 2 tubes at a time, because the alcohol treatment is harsh and might affect germination if is left on seeds too long
 
#Immediately suck off 600 uL of supernatant
 
#Immediately suck off 600 uL of supernatant
#Repeat steps 3 and 4 four times
+
#Repeat steps 3 and 4 four times with water, not ethanol, to wash
 
#Set aside tubes and repeat steps 1 to 5 for all the remaining tubes
 
#Set aside tubes and repeat steps 1 to 5 for all the remaining tubes
 
#Add 200 uL of 20% bleach to each tube
 
#Add 200 uL of 20% bleach to each tube
#Let tubes sit for 10 minutes
+
#Immediatly remove bleach solution
 
#Add 600 uL of water
 
#Add 600 uL of water
 
#Suck off 600 uL of the supernatant  
 
#Suck off 600 uL of the supernatant  
#Repeat steps 9 and 10 four or five times
+
#Repeat steps 9 and 10 four or five times to wash

Latest revision as of 12:14, 1 February 2007

Seed Sterilization

Supplies

  • 70% Ethanol
  • 20% Bleach
  • Sterile Water
  • Seeds

Methods

  1. Put seeds in an Eppendorf tube
  2. Add 200uL of 70% EtOH to tube
  3. Immediately add 600 uL of Water. Do only 2 tubes at a time, because the alcohol treatment is harsh and might affect germination if is left on seeds too long
  4. Immediately suck off 600 uL of supernatant
  5. Repeat steps 3 and 4 four times with water, not ethanol, to wash
  6. Set aside tubes and repeat steps 1 to 5 for all the remaining tubes
  7. Add 200 uL of 20% bleach to each tube
  8. Immediatly remove bleach solution
  9. Add 600 uL of water
  10. Suck off 600 uL of the supernatant
  11. Repeat steps 9 and 10 four or five times to wash