And how to recognize such amplifications on the basis of Tm in the melt curve stage?
As it is a known fact that retropseudo genes have no introns and they can be amplified at cDNA level along with the GOIs.
by definition a retrocopied pseudogene can't be distinguish to the cDNA.
It's also a big problem on the RNA-seq data.
However it's frequent that pseudogenes contain mutations... so it could be possible to design specific primers which recognize or not this mutation.
Eva-green qPCR kit is supposed to be more accurate to separate dissociation peaks with very few mismatch. It could be tested
This paper is likely interesting for you as it describes a commercially-available method to measure and subtract the signal coming from pseudogenes/gDNA: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3326333/
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