Please advise on this.
I have tried all sorts of troubleshooting like-
pH adjustment, ventilation, inversion of the tube gently after adding P2 buffer, overnight culturing time, optimizing the volume, etc.
In the image, the first two lanes show the miniprep extraction of plasmid, and the other lanes show the midiprep - where the genomic DNA contamination is too high.
I'm not sure whether that is really genomic contamination or just a bunch of intercalated plasmids that are knotted up. You could digest with an enzyme that cuts once and see what the gel looks like after that.
However for most applications this is probably clean enough even if there is some genomic DNA, what sort of application are you doing that you think it might be a problem?
Although genomic DNA contamination can be a common issue in Midiprep plasmid extraction, there are various ways to reduce or eliminate such contamination. These methods include selecting bacterial strains with low genomic DNA content, optimizing bacterial culture conditions, using suitable lysis buffers containing detergents, including RNase treatment, purifying plasmid DNA using column-based methods, and measuring the concentration and purity of the extracted DNA. By following these steps, it is possible to obtain high-quality plasmid DNA with minimal genomic DNA contamination.
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