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PCR ready mixes with long shelf lives - (Dec/13/2013 )

Hi all, 

 

Was just wondering, if any body would like to share how commercial PCR mixes manage to have such long storage lives? Given that the Taq is already a part of the mix, it must be quite difficult to maintain its integrity through frequent freeze thaw cycles.

 

Is it just glycerol that gives it a longer shelf life? 

 

Ameya

-Ameya P-

Does your PCR mix not state you should use aliquots to prevent freeze-thawing?

-pito-

Hi Pito, 

 

I am well aware that commercial PCR mixes should be stored in aliquots to prevent frequent freeze and thawing. What I am wondering about is that irrespective of frequent freeze thawing, the PCR mixes work well. I was wondering what ingredients allow these mixes to remain stable over months and over frequent free thaw cycles? 

 

Ameya

-Ameya P-

I guess partly they avoid such problems by having a quite high final amount of Taq. I remember that we used a Qiagen PCR mix that had finally 2.5 U Taq in each reaction (1 x mix). So there's a quite large surplus, and it won't bug the reaction if some portion of the enzyme is dead after some time. But this is not an invitation to freeze thaw such kits as often as possible wink.png ; I'd still use aliquots and freeze-thaw them as rarely as possible.

-hobglobin-

HotStarTaq Master Mix from Qiagen. We just thaw it once (800 ml vial, the rest of the vials are on -20, sometimes even for year or so). It has certified 100% activity after one month on 4 C, but we probably use even older.

I originally made aliquotes, but now everyone is using it, so since we use one vial in +/- month, there is not much reason for doing that anymore. The thawing would probably be bigger problem for the dNTPs.

 

We also have real-time master mixes, some come at bulk 5ml sizes, but they have quite a long fridge shelf life, several months. They don't recommend refreezing.

 

I would say, commercial master mixes are generaly more designed for steady fridge life, that freeze-thaw cycles. And for the user, it's more convenient.

-Trof-