For the first time, I am going to extract plasmid. I would like to know whether gDNA would be extracted during the plasmid extracting? Similarly, is the plasmid extracted when gDNA getting to be extracted?
* The target bacterium is E.coli.
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Herve Levesque
In the classical Birnboim & Doly procedure, gDNA does not renaturate in the cool acid neutralisation step after being denaturate by alcaline conditions. The plasmid do renature due to its small size.
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Laurence Stuart Dawkins-Hall
- Agreed
- Specifically, you denature your gDNA with reagent #2 (SDS/NaOH) and then you precipitate proteins on ice with reagent #3 (3M sodium or potassium acetate pH 4-5) you denature proteins and genomic DNA (but based on topology considerations not your plasmid)
- Thus, when you remove your white floculate you remove your genomic DNA, effectively just leaving plasmid and salt in the supernatant
- Therefore, you will need to perform a second extraction on separate material for genomic DNA
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