I have read that repeated freeze-thawing can damage DNA but then a lot of kit guides recommend freezing for storage. I have stored PCR products and purified PCR products in the fridge and found them to still serve their purpose. Is degradation of DNA more likely at fridge temperature? What's the main difference?
Storage in a fridge will accelerate all kinds of hydrolysis reactions, but for most purposes (meaning subcloning, further PCR, sequencing, analysis by electrophoresis, etc) it doesn't matter. In general, when working with DNA it's far more important to ensure that your samples are nuclease-free than to fret about storage temperature or buffer. I once checked PCR products that had been left on the bench for over a month (room temperature here means 25 to 30oC) and they showed no evidence of degradation; and the performance of the digested plasmid sample I use as a control when doing ligations has remained constant for the last 13 years despite storage in water and multiple freeze/thawing cycles.
That said, storage temperature/buffer does matter when doing quantitative experiments such as qPCR, working with single-stranded DNA, or having to ensure that the ratio of DNA topoisomers in a purified plasmid preparation remains constant (a not unusual requirement when working with DNA vaccines).
This also depends on what your DNA is dissolved in; if in water or buffer (such as TE buffer), DNA is very stable when frozen. If kept refrigerated, DNA can degrade in water whereas it will not degrade in TE under the same conditions. Water can cause degradation of DNA by hydrolysis at 4 degrees, whereas this appears not to be the case when kept frozen.
Despite the above, I don't think you run the risk of your PCR products becoming unusable if they are stored in water in the fridge - unless they are there for years! As for freeze-thawing, I have never had DNA become problematic even when I freeze-thaw it multiple times.
Check out the attachment.
It all depends on what you want to do with your PCR products and long term storage is not recommended for PCR products.
Freezing will not affect the DNA. Since this is not circular DNA, negligible amount DNA will be degraded.
Hi Emma,
Freezing does not affect the DNA as DNA is quite stable entity. However, quick freezing and thawing may affect it which may cause the formation of single stranded DNA. This not due to degradation but due to random annealing of the strands. This is particularly important when you have to clone the PCR fragment.
I have used both, refrigerator and freezer without any problem. However I have not store them for a long time.
Storage in a fridge will accelerate all kinds of hydrolysis reactions, but for most purposes (meaning subcloning, further PCR, sequencing, analysis by electrophoresis, etc) it doesn't matter. In general, when working with DNA it's far more important to ensure that your samples are nuclease-free than to fret about storage temperature or buffer. I once checked PCR products that had been left on the bench for over a month (room temperature here means 25 to 30oC) and they showed no evidence of degradation; and the performance of the digested plasmid sample I use as a control when doing ligations has remained constant for the last 13 years despite storage in water and multiple freeze/thawing cycles.
That said, storage temperature/buffer does matter when doing quantitative experiments such as qPCR, working with single-stranded DNA, or having to ensure that the ratio of DNA topoisomers in a purified plasmid preparation remains constant (a not unusual requirement when working with DNA vaccines).
From my experience I would say, better store it in a freezer. In normal cases its ok to store it in fridge or as Martin says you can keep it on your table at room temperature (but not for long). But if you have a higher concentration of MgCl2 in your PCR reaction then for long term storage you have to use a -2o deg C freezer.
Thanks for all the answers! To date, samples stored in the fridge seem to be working fine. I think I will store them short-term in the fridge and long-term in the freezer from now on.
to store PCR products use10/0.1 TE [10mM Tris, 0.1 mM EDTA, pH 8.5] . The tris helps stop the solution from becoming acidic (cause by CO2 being absorbed from the air) and the small amount of EDTA doesn't interfere with the sequencing reaction.
You have store pcr products at -20 freezer if you are using it for cloning or sequening
Dear all
I stored a multiplex PCR product (DNA) for about a month at +4c , then when I try to measure the concentration using quite, it is totally out of limit( high concentration which the machine can't read). NB it was in normal at the very begging when I put it there. So do any one share me his experience when the concentration increase as such or what the solution could be
Best regards,
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