My qPCR taqman reaction has stopped working. It's a duplex reaction that amplifies a sample and an internal control having FAM and VIC probes respectively. I use a Quantstudio 6 flex equipment. Everything was fine until recently having no or weak amplifications. I repeated the assays with positive controls and getting no or weak amplification curves. I have prepared new primer stocks and all other reagents are fresh. Any thoughts?
If everything is fine with the reagents the equipment detector could be still wrong, so I would test that as well with an old plate/tubes (if you still have them). You should be able to read out high copy numbers from those instantly if the qPCR machine is fine.
If your PCR reaction worked, you should be able to visualize the product of your PCR with an agarose gel.
Ajith Kumar nothing changed really. Only a new batch of dNTPs and taq polymerase but they are from the same supplier.
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