Antisense PCR

Brisco and Morley described a method whereby antisense oligonucleotides can be used to switch off the activity of primers at a defined point during the PCR. This led to the development of a highly efficient nested qPCR which could be performed in a closed tube. In this nested PCR, a decrease in the annealing temperature enabled the antisense oligonucleotides to hybridise to and inactivate the outer primers, and at the same time enabled the inner primers to hybridise and extend. Since both outer and inner primers were at their optimal concentration for annealing, the reaction was as efficient as a classical one-round PCR. More recent studies have shown that primers which are inactive can be activated during the PCR using the same approach.

Brisco and Morley suggested that, in addition to performing nested PCR in a single tube, antisense PCR might be useful in quantifying rare targets, when non-specificity might be a problem, and in performing multiplex PCR. Recent studies indicate that antisense PCR is useful in providing single-stranded DNA.

See also Setting up an antisense PCR.

Reference

Brisco, M.J., Bartley, P.A., and Morley, A.A. Antisense PCR: A simple and robust method for performing nested single-tube PCR. Analytical Biochemistry 409:176–182 (2011)