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weird DNA bands in the Minipreps; competent cells contamination? - (Oct/19/2006 )

Hi all

I am having troubles with the minipreps. I've ligated a PCR product (blunt ended) with a Zero Blunt TOPO vector. I've transformed DH5 alpha competent cells with the ligation and I get the same weird band (1,5Kb size) in all digestion (even if I digest it with different enzimes I get the same band). Somebody in the lab is having the same strange band. I haven't seen this before; Could it be a phage contaminating our DH5 alpha cells? We are gonna try other different competent cells. Does anyone have had a similar problem? I haven't used the TOPO vector before; does anyone have had problems with this type of ligation?

Thanks a lot in advance.



Could it be super-coiled plasmid that did not get digested?


could u show a pic of the gel. what sizes are the vector and insert?

i wonder if due to some reason the enzymes r unable to digest the DNA.


How are you performing your mini-preps? Are you using any kit of doing it with homemade solutions?
maybe the contamination band is bacterial DNA. This can occur if you are not gently when you add you second solution (commolly refered to as the P2) that breaks up the bacterial cell...

The contamination band is normally and upper band though (not at 1.5kb!!).