I have noticed that the aliquoted diluted dNTPs as they age stop giving results for PCR. Do anyone know the reason for this or have experienced the same? For some reason I think that they retain their properties only when they are concentrated.
Hi, you are absolutely right they are stable at conc like 100mM I make my stock solutions with this conc. In my opinion the reason is the lost of the phosphate and instead of dNTP you finaly have dNDP
Hi, you are absolutely right they are stable at conc like 100mM I make my stock solutions with this conc. In my opinion the reason is the lost of the phosphate and instead of dNTP you finaly have dNDP
The dNTPs are sensitive to hydrolysis on the phosphate groups. Repeated freezing and thawing cycles of your stock promote this. So make aliquots of them, which are not too big to be used up in a relative short time.
A hypothesis is as dNTPS start breaking down they mimic nucleoside analogs that are known to cause termination of replication. Compounds likes acyclovir.
dNTPs may decompose into nucleoside di- and mono- phosphates via a disproportionation reaction. It is advisable to keep nucleotides frozen in aliquots and avoid multiple freeze/thaw cycles. Additionally, you may keep dNTP solution on ice once thawed to minimize degradation.
Because dNTPs are sensitive to hydrolysis on the phosphate groups. sothat it breaks Nucleloside analog molecule..that will result termination of replication.
I have experience to use new TaqPol with good DNA template with old dNTPs and somehow not get results. I have doubt on my old dNTPs. However I can only said firmly when I try the same reaction with same condition with new dNTPs.
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