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> | ||
| + | <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> | + | <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.