generally annealing temperatures will be just 3-4 C below Tm of
primers. if the melting point of primers is very high like you said in
the range of 90 , at that hight tempertaure all double stranded dna
will split off and so annealing will not take place., because
annealing happens only in the range of 50 - 65 C. hope i'm correct..
if i'm wrong anyone can correct me...
On Jan 27, 9:43 pm, Jonathan Nesser <jonathan.nes...@gmail.com> wrote:
> Sorry to start yet another new thread for such a small question, but I
> don't want to pollute other threads with off topic posts... I've been
> trying to understand why primer melting temperatures have to be in the
> 55-65C range when PCR cycles call for 94C for DNA melting... I've tried to
> find an explanation for this but haven't been able to... Wouldn't that
> result in the primers splitting off from the template DNA before you reach
> the extension cycle of 75C (or around there)? I'm sure I'm wrong because
> obviously these PCR programs work, but I don't know WHY I'm wrong, and
> would like to understand. Thanks again for explaining something that is
> probably so common knowledge that nobody feels the need to address it in
> writing :)
>
> Jonathan Nesser
> jonathan.nes...@gmail.com
> diybioandneurosci.blogspot.com
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