"Ghost" bands in plasmid preps?

[Ned Mantei]

In article <B461CBA9.C4E%Simon.Dawson at nottingham.ac.uk>, Simon Dawson
<Simon.Dawson at nottingham.ac.uk> wrote:

> Hi all,
>         Does anyone ever see what I can only call "ghost' bands if they
> over-expose an ethidium bromide stained agarose gel of plasmid DNA? They
> usually appear to run somewhat faster than what I would think is the
> supercoiled DNA band and you only seem to see them if you overexpose the
> gel.

More or less permanently denatured supercoiled DNA, resulting from the
alkali treatment. During denaturation the strands can't separate, so every
turn that the helix unwinds must be compensated by a reverse turn of the
molecule as a whole. This can lead to such a snarl that the DNA can't
rewind in a reasonable time. This is also why plasmid DNA is heated in
alkali to prepare templates for sequencing.

-- 
Ned Mantei
Dept. of Cell Biology, Swiss Federal Institute of Technology
CH-8093 Zurich, Switzerland