Difference between revisions of "Ethanol Precipitation of Vector DNA"

From GcatWiki
Jump to: navigation, search
 
Line 1: Line 1:
 
# After digestion with restriction enzymes (ex. Normally a 40 µL digestion)
 
# After digestion with restriction enzymes (ex. Normally a 40 µL digestion)
# Add 1/10 the digestion volume of 3M NaOAc pH 5.2 (ex. 4 µL of NaOAc)
+
# Increase volume to 90 µL with dH20
# Add 2X the reaction volume of ethanol (ex. 88 µL of ethanol)
+
# Add 10 µL of 3M NaOAc pH 5.2
 +
# Add 2X the volume of ethanol (ex. 200 µL of ethanol)
 
# Vortex and put in -80°C freezer for 15 minutes
 
# Vortex and put in -80°C freezer for 15 minutes
# Centrifuge for 15 minutes (Place the tube hinge out in the microcentrifuge to know where the pellet is.
+
# Centrifuge for 30 minutes (Place the tube hinge out in the microcentrifuge to know where the pellet is)
# Dump off the liquid and allow to dry, either sitting on the bench or with the lyophilizer.
+
# Dump off the liquid and allow to dry, either sitting on the bench or with the lyophilizer

Latest revision as of 12:53, 4 June 2015

  1. After digestion with restriction enzymes (ex. Normally a 40 µL digestion)
  2. Increase volume to 90 µL with dH20
  3. Add 10 µL of 3M NaOAc pH 5.2
  4. Add 2X the volume of ethanol (ex. 200 µL of ethanol)
  5. Vortex and put in -80°C freezer for 15 minutes
  6. Centrifuge for 30 minutes (Place the tube hinge out in the microcentrifuge to know where the pellet is)
  7. Dump off the liquid and allow to dry, either sitting on the bench or with the lyophilizer