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Different primer concentration in qPCR - (Feb/25/2014 )



It is the first time I am using these three set of primers, I would like to optimize the primer concentration beforehand.


My question is what if the optimal concentration between these sets of primers differs, are we allowed to run qPCR with different primer concn?


For example:

Primer A--- Forward 150nM, Reverse 300nM

Primer B--- Forward 300nM, Reverse 300nM

Primer C--- Forward 300nM, Reverse 150nM





Are you running quantitative (real-time) PCR with three sets of primers, each one separately?


If so, it doesn't matter. Each reaction has it's own standards (absolute or relative) to which the results are in relation.