Annealing palindromic linkers

[Peter Ashby]

In article <3a63fb84$1 at news.unibe.ch>, "Bruno Cenni" 
<bruno.removethis.cenni at removethis.insel.ch> wrote:

> Hi all,
> 
> we're (expectedly) having troubles cloning a linker into a plasmid. This
> linker consists of two complementary 33 bp oligos that after annealing 
> have
> open restriction enzyme sites at their ends. The problems comes probably
> from the fact that the oligos contain two complementary inverted 6 bp
> recognition sites thus giving a palindrome (spaced by 3 bp). Any 
> suggestions
> on conditions to increase the proper annealing of the oligos without too
> much of the self-annealing and stem-loop formation?

Are your oligos phosphorylated? it makes quite a difference when doing 
this and I would suspect that before assuming your oligos haven't 
annealed. A cheap way to achieve this is to make your oligos overlapping 
and then cut them after annealing.

Peter

-- 
Peter Ashby
Wellcome Trust Biocentre
University of Dundee
Dundee, Scotland
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