Poor PCR or qPCR efficiency? It can happen to everyone, even to an expert. One thing we all learn over the years about the PCR – you shall be always awake and ready for optimization and changes.

What do you usually suspect as a PCR problem number one? All PCR trouble shootings speak the same language: low PCR yield or no PCR product can be a result of different suboptimal PCR reaction parameters.

Starting from the core, one could suspect the template. DNA or cDNA can be impure, can be damaged, and can be used in too high amount or at too low concentration. Same about the primers; they might be designed wrong, synthesized and purified inappropriately or may contain contaminants that inhibit the PCR. PCR reaction conditions such as the concentration of the buffer components, dNTPs, dropped PCR polymerase activity or wrong cycling parameters may also result in too low amplification yield.

There are numerous articles and even books written about the PCR problematic. But the rule of the thumb is to set up the right control reactions and to changereaction parameters one by one. It may happen that you have to change the Taq Polymerase or dNTPs, or qPCR master mix.  

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