View source for TOPO Cloning

#PCR desired insert using Pfu polymerase.  Add 1 uL of Taq and mix.  Incubate 10min at 72C to add A to 3’ ends.  Use immediately
#If PCR product is a single band, no need to purify further.  If several bands, gel purify.
#Combine 4 uL PCR product, 1 uL Salt Solution, and 1 uL TOPO vector.  Mix gently
#Incubate at room temperature for 5 min, place on ice.
#Thaw tube of OneShot cells on ice
#Add 2 uL reaction and incubate on ice 5-30 min
#Heat shock 30s at 42C without shaking
#Place on ice for 2 min
#Add 250 uL of room temperature SOC media
#Incubate at 37C for 1h
#Plate 50 uL on plate and grow overnight