Awesome Double Zap Back Transformation

Guy Oshiro oshiro_G at defiance.hsc.colorado.edu
Tue Apr 4 13:48:02 EST 1995


Hey yeast-netters,

Our lab just tried out an awesome back transformation protocol from the
Brent lab.  Check it out on their web site.

They basically do a double zap transformation.
1. Prepare your elctrocompetent e. coli (i.e. DH5alpha)
2. Mix 80 ul of the e. coli cells with an equal amount of a yeast
colony picked straight off of a plate.
3.  Transfer to a electroporation cuvette.
4.  Zap them at 1500 v to zap the plasmid out of the yeast.
5.  Chill on ice
6.  Zap again at 2500 v to zap the plasmid into the e. coli.
7.  Recover in LB
8.  Plate out onto L+amp plates.

It works and it is very simple.
You can get more sophisticated and select for a single plasmid if there
are multiple plasmids inside the yeast by using a Pyr F e. coli strain.
 And selecting on a drop out plate.
 
Guy Oshiro
EMAIL: Oshiro_G at defiance.hsc.colorado.edu

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