could anyone tell me if we standarize primer concentration(0.1-0.5uM), then which we should use for further qpCR analysis if we are getting no primer dimer in either case, ideally one should take lowest primer concentration, what I am getting different fold change with different primer concentration. Which one to believe??
Dear Shilpi
If you are not getting primer dimer then you should use higher concentration because sometime lesser amount of Primer can be a limiting factor for good amplification and higher amount of primer can be a cause of noise (so the working concentration should be at ~0.05 uM/reaction.
All the best
I think Abishek meant 0.5 uM (500 nM). And I agree: use the higher concentration here.
I second the 0.5 uM but with one caveat: use the primer concentration recommended with your Polymerase mastermix.
For example, the Qiagen Mix I use recommends 1uM primer each.
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