Calculating Tm of PCR Product for RT-qPCR Melting Curve

[mountain]

I know how to calculate Tm of a primer but what about of a PCR product?

I want to run the melting curve in RT-qPCR to test for the products and i want to know Tm of the products to be sure.


Thanks.

 

[Monique]

Why not use the same calculation as the Tm of a primer? A good calculator should takes into account the length of the nucleic acid. How do you calculate it now?

 

[Monique]

Here's an online calculator http://biotools.idtdna.com/analyzer/

 

[mountain]

Why not use the same calculation as the Tm of a primer? A good calculator should takes into account the length of the nucleic acid. How do you calculate it now?

The difference between a primer and a PCR product is that the PCR product is a double stranded DNA while the primer is just a singel strand. So should i calculate Tm for the PCR product like as it is a single strand?

 

[iansmith]

Although you calculate Tm with a single strand for primer, it is use to separate your double stranded DNA.

So for your PCR product calculate with the single stranded DNA sequence.