Glycerol inhibition of PCR?

Mario Vaneechoutte Mario.Vaneechoutte at rug.ac.be
Fri Feb 2 02:31:49 EST 1996


Since more than a year we add glycerol (10%) and cresolred to our PCR 
mixture (according to Pomp, D., and J. F. Medrano. 1991. Organic solvents
as facilitators of polymerase chain reactions. BioTechniques 10:58-59). 
One of the advantages is that we save a pipetting step after PCR. 
Glycerol and cresolred can be used as a PCR compatible gel loading 
solution, so we don't have to add gel loading solution after PCR.

Things worked fine, until recently. Omitting glycerol and cresol red  
resulted in good amplifications again.
Now we compared PCR and PCR with 10% glycerol and found the same 
glycerol used before to be inhibitory. Glycerol was stored as a 50% 
solution, aliquotted, at room temperature (in sunlight). Making new 
50% solution from the original bottle showed that also this freshly 
made solution clearly inhibited in comparison with the control PCR 
(although less than the old aliquots, which inhibited completely).

Since glycerol is known (in all hand books) to enhance PCR (e.g. by 
protection of the 'thermostabile' polymerases at high temperatures), 
something which we have been able to confirm, this is a peculiar finding.
I wonder what can change in glycerol during storage such that it becomes 
inhibitory?
We'll try new bought glycerol soon, but maybe someone has some 
suggestions already.







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