Is it possible to accurately and reliably estimate amplicon length or amplicon length distribution from qPCR melting curve/dissociation curve? Are there any articles on this?
If it matters, my purpose is to estimate mean fragment length of illumina libraries from qPCRs without having to run a Bioanalyzer or Tapestation.
Hi,
I think, that this approach is not ideal. Melting curve is based on dissociation characteristics of double-stranded DNA during heating (base pairing not the length of DNA).
If you need to check your library without Bioanalyzer or Tapestation, you can run gel-electrophoresis where you should see equally distributed smear.
It is also good to measure the concentration of a library on Qubit.
It is not possible to determine the length of the amplicons because the dissociation curve depends on the quantity of the gene and this has nothing to do with the length of it
No, you can't determine amplicon length by melt temperature. The melting temperature is a combination of length and GC content.
Run out your sample on an agarose or acrylamide gel to determine fragment length
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