Difference between revisions of "Transformation via Electroporation"
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| − | -Set the electroporation apparatus to 1 | + | -Set the electroporation apparatus to 1.8 kV, 25 microfarads, and the pulse controller to 200 omega |
-Add 1.5 microliters of DNA to be transformed to 40 microliters of electrocompetent cells on ice and mix | -Add 1.5 microliters of DNA to be transformed to 40 microliters of electrocompetent cells on ice and mix | ||
Revision as of 14:17, 20 July 2015
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Contents
Purpose
This protocol is used to transform electrocompetent cells with a plasmid.
The electrocompetent cells are made using the protocol found on Preparation of Competent Cells for Electroporation.
Materials Needed
-Electroporation apparatus
-1 mm electroporation cuvette
-micropipette
-LB broth
-Electrocompetent cells
-DNA
-microfuge tubes
-ice
-kimwipes
-LB or NBY agar plates
-28 C incubator
Protocol
-Set the electroporation apparatus to 1.8 kV, 25 microfarads, and the pulse controller to 200 omega
-Add 1.5 microliters of DNA to be transformed to 40 microliters of electrocompetent cells on ice and mix
-Transfer the DNA and cells to a pre-chilled 1 mm electroporation cuvette
-Wipe the cuvette with a kimwipe and place it into the sample chamber
-Deliver a pulse by pushing in charge button
-Remove the cuvette and ad 1 mL LB broth to the cuvette
-Transfer the broth and cells to a sterile tube and incubate for 2 hours at 28 C with shaking
-Place aliquots of the mixture on LB or NBY agar plates with the appropriate antibiotics for the cells
-Incubate at 28 C for 2 days
Notes
Papers where this or a similar method has been used
Related Ontology Terms
References
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