What factors affect melt peak [-d(RFU)/dT) in real-time PCR?
What should I do if I want to increase my melt peak [-d(RFU)/dT)?
The melt curve is the LEAST important piece of information you get from qPCR.
Unless you are using melt-curve-based genotyping (which is a separate technique that answers different questions), then don't bother worrying about this.
You get relatively taller peaks when you have relatively more product. Kind of. But since it's RELATIVE, this information is honestly not useful.
Real-Time PCR (Probe type) melt peak factor, also known as the Tm (melting temperature) or Tm peak, is a measure of the stability of a double-stranded DNA molecule. It is the temperature at which half of the double-stranded DNA dissociates into single strands. In real-time PCR, the Tm peak factor is used to determine the specificity of the PCR reaction.
The Tm peak factor is used to determine the specificity of the PCR reaction because it is affected by the length and nucleotide sequence of the DNA fragment, as well as the buffer conditions and the presence of other DNA sequences in the reaction mixture. A specific and efficient PCR reaction will produce a single, sharp Tm peak, while a non-specific reaction will produce multiple, broad Tm peaks. The Tm peak factor is therefore a useful tool for assessing the specificity and efficiency of real-time PCR reactions.
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