Difference between revisions of "Team:Birkbeck/Electroporation"

 
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<h1>Transformation of plasmids into electrocompetent cells</h1>
 
<h1>Transformation of plasmids into electrocompetent cells</h1>
  
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<p>For more details about making electrocompetent cells, please see our <a href="https://2015.igem.org/Team:Birkbeck/Electro-Competent_E._coli_Cell_Protocol">Making electrocompetent <i>E.coli</i> cells protocol</a>.</p>
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<h2><b>Electroporation protocol.</b></h2>
 
<h2><b>Electroporation protocol.</b></h2>
 
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<a href="https://2015.igem.org/Team:Birkbeck/Experiments">Back to experiments and protocols</a>  
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<li><a href="https://2015.igem.org/Team:Birkbeck/Experiments">Back to experiments and protocols</a></li>
 
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Latest revision as of 17:35, 18 September 2015

Transformation of plasmids into electrocompetent cells

For more details about making electrocompetent cells, please see our Making electrocompetent E.coli cells protocol.


Electroporation protocol.

  • 1. Thaw out the 50 µL cell suspension aliquot on ice for ca. 30 minutes. Concurrently, cool electroporation tubes on ice.
  • 2. Add 100pg-1µg of plasmid DNA to the cell suspension (approximately 1-5 µL of plasmid solution).
  • 3. Mix plasmid and cells thoroughly and continue to incubate on ice for 5-10 minutes.
  • 4. Transfer the mixture of cells and plasmids to an electroporation tube.
  • 5. Electroporate cells; for E.coli, this is done at 2.5 kV, 200 Ohms, 25 uF.
  • 6. Immediately add 450-950 uL of LB broth.
  • 7. Incubate tubes at 37oC for 60-90 minutes in a statis incubator.
  • 8. Plate 100 uL of cells onto agar plates with appropriate antibiotic resistance.
  • 9. Incubate overnnight at 37oC in a static incubator.

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