Gold Nanoparticle Effects in Polymerase Chain Reaction: Favoring of Smaller Products by Polymerase Adsorption

Abstract

Gold nanoparticles were recently reported to reduce the formation of nonspecific products in polymerase chain reaction (PCR) at remarkably low temperatures, with hypothesized mechanisms including adsorption of DNA and heat-transfer enhancement. In contrast to these reports, we report that gold nanoparticles do not enhance the specificity of PCR but rather suppress the amplification of longer products while favoring amplification of shorter products, independent of specificity. Gold nanoparticles bearing a self-assembled monolayer of hexadecanethiol did not affect PCR, suggesting that surface interactions play an essential role. This role was further confirmed by experiments in which a similar effect on PCR was observed for the same total surface area of particles over a 100-fold range of per-particle surface area. The effect was seen with Taq and Tfl polymerases but not with Vent polymerase, and the effects of nanoparticles can be reversed by increasing the polymerase concentration or by adding bovine serum albumin (BSA). Transient high-temperature nanoparticle pre-exposure of PCR mix containing polymerase but not template or primers, followed by nanoparticle removal, modified subsequent nanoparticle-free PCR. Interaction between polymerase and gold nanoparticles was confirmed by changes in nanoparticle absorption spectrum and electrophoretic mobility in the presence of polymerase. Taken together, these results suggest that the nanoparticles nonspecifically adsorb polymerase, thus effectively reducing polymerase concentration.