Ghost bands on plasmid preps

Claudia Sutton cas9 at cornell.edu
Wed Apr 4 08:08:06 EST 2001


In article <3ACB036B.6B1CD135 at cmm.ki.se>, Eric DUFOUR
<Eric.Dufour at cmm.ki.se> wrote:

> I just found that explanation in the Quiagen plasmid purification handbook :
> Extra DNA bands on analytical gel -a Dimer form of plasmid -b Plasmid has
> formed denatured supercoils "this species run faster than closed circular and
> are resistant to restriction digest ". Since their protocol is based on
> alcaline lysis... They also state as it as been said before that it is due to
> too long incubation in NaOH/SDS (too long lysis). I think there is a comment
> on that in maniatis also...
> 
> Eric Dufour

I discovered the same thing years ago, the band is denatured DNA. In my
case it was plasmid+insert-dependent (one clone gave me ONLY denatured
form). It can be almost completely solved by decreasing [NaOH] to 0.15M in
solution 2.

Claudia




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