Glycerol inhibition of PCR?

[Mario Vaneechoutte]

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.

[Elliott Dawson]

We too have found that glycerol can enhance some PCR reactions and the
combo with dye is certainly convenient. However,there are certain caveats
with respect to its use.
1. We generally titrate glycerol over a range of 1-10%
2. We have found that in some cases %'s lower than 10 are best, i.e. 2%
3. In other cases we find that glycerol is of no benefit and is detrimental
4. Glycerol is subject to oxidative degradation and we store our 50% stock
at -20C.
5. Glcerol may or may not be benificial depending on primer design, target
complexity, type of polymerase buffer, volume, etc.
6. If you are still in need of a cosolvent then try DDMSO over the range of
1-10%.