I want to transfect Adipose Derived stem cells using a clone of my own. My problem is that I get a lot of contamination (not only genomic, but due to the nature of the technique, I get undesired products I haven't been able to get rid of).

So I was thinking about running a gel with my plasmid DNA samples and gel purify my desired bands. I don't know, however, if Ethidium Bromide contamination in my sample would be a problem or if a regular gel extraction kit gets rid of it.

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    $\begingroup$ No, there is absolutely no problem with that. And you get rid of the EtBr in the purification anyway. $\endgroup$
    – Chris
    Jul 1 '15 at 5:53
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    $\begingroup$ Ethidium toxicity is probably not as bad as they make you think, they use it at like 1 mg/kg in cattle to prevent parasite infections. $\endgroup$
    – user137
    Jul 1 '15 at 5:58
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    $\begingroup$ And I should point out that it's not efficient to use plasmid purified from gel directly in a mammalian cell transfection. You should consider a larger scale prep. If its a problem of incorrect plasmids, transform your plasmid mix in bacteria, pick several SINGLE colonies and miniprep them, sequence and try again. $\endgroup$
    – user137
    Jul 1 '15 at 6:01
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    $\begingroup$ Possible duplicate of biology.stackexchange.com/questions/14047/… $\endgroup$
    Jul 1 '15 at 7:20
  • $\begingroup$ Thanks for your insightful answers! To answer user137: I cannot expand my plasmid in bacteria to use a larger prep since the OriR sequence is excised and degraded from the construct after induction with arabinose. This is what I'm trying to get: ncbi.nlm.nih.gov/pmc/articles/PMC4144359 Again, thanks $\endgroup$
    – Varekai
    Jul 1 '15 at 17:04

Purification and isolation of DNA bands by cutting them from agarose gel is commonplace (Lee et al., 2012). The purification step after excision of the band gets rid of most of the EtBr and other impurities. E.g., see the websites from Isogen and Sigma-Aldrich.

- Lee et al. J Vis Exp (2012); 62: e3923


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