Pair of primers (SYBR) with more than one amplicon product(?)
Posted 27 March 2013 - 04:11 AM
I have a question.
I am trying to design a pair of primers in a repetitive sequence, but i am facing the issue below:
The forward primer is ok and align in only one site in the sequence.
But the reverse primer align in four sites downstream the sequence amplifying 4 amplicons: 213bp, 425bp, 637bp and 849bp.
My question is: at the end of PCR, how many amplicons will I have?
Theoretically I know that I will have 4 amplicons, but I was thinking if the first aligment of the reserve primer (213bp) will allow the other bigger amplicons to be produced.
What do you think about it?
Posted 27 March 2013 - 04:15 AM
Posted 27 March 2013 - 06:20 AM
does the second primer aneal to the sequence that is 100% complementary (in all 4 cases) ?
It is a repetitive sequence, so the primer align perfectly in all 4 downstream sequence.
Posted 27 March 2013 - 06:54 AM
There will be the most of shorter fragments and least the longest ones.
I haven't tried such experiment yet, but time will come.
Posted 28 March 2013 - 03:22 AM