Ghost bands on plasmid preps

Nick Theodorakis nicholas_theodorakis at urmc.rochester.edu
Sat Mar 31 23:11:06 EST 2001


In <3ac67107_2 at news01.one.net.au>, Daniel MacArthur wrote:
>
>Dear group,
>
>I'm an undergraduate biochemistry student currently attempting to write up a
>practical report on a plasmid miniprep practical. Briefly, we took two E.
>coli colonies transfected with pBluescript IISK+ vectors (one containing a
>fragment from an EcoRI digest of lambda phage DNA and one with no insert),
>purified the plasmids using a Quantum Miniprep from Bio-Rad, digested the
>plasmid DNA using EcoRI, BamHI, and a combination of the two, and then ran
>out the products on an agarose gel.
>
>The experiment worked beautifully, but I'm having a little trouble with
>interpreting the results. In several of the lanes there are faint bands
>running at roughly half the size of the full-length linear plasmid, which my
>demonstrator called "ghost bands". 
[...]

Hengen's denatured ghost bands sounds like a reasonable explanation to me. Try
reducing the amount of time during lysis in the alkali/SDS step. The times that
I've seen them has been when I've been doing a lot of minipreps and they
probably spent too much time in the alkali.

Nick


-- 
Nick Theodorakis
nicholas_theodorakis at urmc.rochester.edu




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