Tricine buffer for PCR

Peter Gegenheimer peterg at rnaworld.bio.ukans.edu
Mon Dec 19 18:39:00 EST 1994


In <3crhti$g70 at highway.LeidenUniv.nl>, jasper%ruly46.leidenuniv.nl (Jasper Saris) writes:
>Michael Cooley (szcooley at chip.ucdavis.edu) wrote:
>: I hear tell of a buffer for PCR based on Tricine. It's supposed to hold a 
>: higher pH at the elevated temperatures of the PCR. Has anyone tried this 
>: and does anyone know the recipe for making it?
>
>
>: #%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%
>: %                                                        #
>: #   Dr. Michael Cooley         Practice Random Kindness  %
>: %   mbcooley at ucdavis.edu    and Senseless Acts of Beauty #
>: #                                                        %
>: %#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#%#
>
>Hoi Michael,
>
>We use TRIS.HCL at pH8.4, pH8.9 or pH10 on roomtemperature. Most PCR sets 
>will work in all three buffers. Often the pH10 variant performs better 
>for smeary PCR's. The idea behind this is a higher annealing 
>(and elongation?) specificity of the several "Taq" polymerases we use.
>Normal is 50mM KCl, 10mM TRIS, 0.5-2.5mM MgCl.
>
>I hope this is helpfull information,
>
>Jasper Saris
>
Thanks for the grins! "Tris-HCl, pH 10" is another one of those weird urban 
myths. Since the pH of pure Tris base is about 9 ( at 1 or 2 M), it will be a 
humorous task to reduce it to pH 10. Also, of course, Messrs. H & H will spin in
their graves at the thought that such a beast could serve as a buffer...
(Ask if you don't know Messrs. H & H. Just doing my 2nd job as a Biochem prof!)
PAG.
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