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Select the optimal primer-concentration

real-time pcr primers concentration optimization

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#1 whitelines

whitelines

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Posted 15 February 2013 - 05:36 AM

Hi,

How do I decide the optimal primer-concentration for my assay? Available litterature says run a matrix of concentrations and choose the one with lowest Ct-value.

However I am concerned that the more substantial formation of primer-dimers at the concentration giving the lowest Ct (red arrows in figure) might have an impact on the sensitivity of the assay when working with lower concentrations of template?
(My goal is to procude the most sensitive assay possible (viral DNA). Primers are published and used by others.)

Figure:
Primer koncentration = 300nM (red arrows)
Primer concentration = 100nM (blue arrows)

Hope I could have som feedback on this oneprimer tes.jpg primer tes.jpg :)
Thanks in advance.





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