Monday, 29 April 2019

pcr - When designing primers how important is the GC clamp?


I'm designing a set of primers and reading about the principles of primer design one of which is:




GC Clamp: The presence of G or C bases within the last five bases from the 3' end of primers (GC clamp) helps promote specific binding at the 3' end due to the stronger bonding of G and C bases. More than 3 G's or C's should be avoided in the last 5 bases at the 3' end of the primer.



From here.


My question is how essential is it to have a GC clamp?



Answer



It's hard to provide an objective answer. If you have a decent length and good complexity, even a single terminal 3' G or C would do. Of course, one has to take into account the primer's overall GC:AT ratio and things like annealing temperature.


Here's a link to diverse opinions on the topic and it has this nugget (which I subscribe to when possible):



FWIW, my preferred offerings to the PCR gods are primers with a single G or C 3', FWIW. Seems to keep 'em happy most phases of the moon.




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