Weird kinased oligo pattern...
John E. Fox
altabios at bham.ac.uk
Tue Jun 7 06:26:12 EST 1994
In article <Cr0uCD.Hsp at midge.bath.ac.uk>, bssgklm at midge.bath.ac.uk (G K L Munro) says:
>I have been having some trouble with oligos which I am using for PCR and
>sequencing. Finally last week I kinased them up to have a look (Iknow, I
>should have done it straight away!). Anyway, although there is a reasonably
>sized band at the right size for the oligo, there are ladders extending in
>each direction!!! The oligos are 19mer, 22mer and a 25mer. They were
>kinased and run on a 20%acrylamide/7M Urea gel at 50C. I have never seen a
>ladder extending in the n+1 direction before. How does this happen?
I experienced similar problems some time ago. Although it was mainly in the n-1 direction.
One of my customers had kinased an oligo to check its purity. It was a complete ladder from
10 up to 50, the target and then several bands beyond.
We remade the oligo only to get the same results. All the synthesis QC looked OK.
Eventually the customer did a UV shadow and saw a perfectly clean oligo.
The conclusion was that the kinase was degrading the oligo, although I am still not
sure where the n+1 bands came from.
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