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Primers have worked well but now getting primer dimers?

Bisulfite PCR PCR Bisulfite converted DNA Primers Primer Dimers

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#1 Epigeneticist

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Posted 28 January 2014 - 10:34 AM

I have some bisulfite PCR primers. They had worked well for us in the past (numerous years and been published). Recently I am getting primer dimer formation from them. I even made fresh dilutions from my stock tube. Old working and new working primer both give primer dimers. I am not using a hot start polymerase but never did in the past and they worked fine. Can primer degrade and give rise to primer dimer? This happened before and was corrected after I made fresh working primers. Maybe my stock is degraded. It's not an issue with optimization. I am worried about a contaminate or degradation of my primers. Suggestions? Comments?

 

Thanks!



#2 Epigeneticist

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Posted 30 January 2014 - 09:28 PM

Problem doesn't seem to be due to primer degradation or poor sample quality/quantity. I repeated the PCR with fresh working primers from the same stock solution but replaced all other reagents and consumables. PCR worked for all samples and the product was very clean and strong. Probably due to some contaminant - maybe primer dimer contaminated the water, reagent or consumables. 



#3 Golgicik

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Posted 03 February 2014 - 01:40 PM

I would just order same primers again and change other solutions, water, buffer etc. 







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