So I have been reading the principles of qPCR for the amplification of DNA. So the reaction takes place after the primers have been combined to DNA single strands, and enzymatic reactions elongate the primers.
My question is, how does the reaction come to a stop? Do the primers just keep getting longer and longer?
Thanks a lot for your feedbacks.
in the very first cycle of pcr the dna is made single stranded by heating then the primers anneal on cooling and extension takes place. This first extension runs for a large distance from the primer and only stops when the enzyme falls off the now double stranded product. So at the end of the first cycle you have many molecules of random long lengths.
At the second cycle the reverse primer anneals in its place on the long amplimers and extends but it cannot extend a long distance because the forward primer defined one end of the amplimer so you cannot extend past that end.
After the second cycle there is still genomic template being extended to random lengths but the product between the 2 primers is doubled every cycle while the genomic target is a fixed amount of dna so after 10 cycles there will be 1000 times as much amplimer as template and after 20 cycles 1000000 times as much amplimer as template so the small template contribution to the pcr product can be ignored as being very small
It's the same as in standard PCR. This video might help!
https://www.youtube.com/watch?v=iQsu3Kz9NYo
- Badges
- Science method