Mechanism of the interaction between Au nanoparticles and polymerase in nanoparticle PCR

Abstract

Nanoparticle PCR is a novel method to optimize DNA amplification. It performs well in improving specificity, enhancing sensitivity and speed. Several mechanisms were proposed in previous studies: one was based on the interaction between gold nanoparticles (AuNPs) and DNA while the other was attributed to the heat transfer property of AuNPs. In this paper, we propose that the interaction between AuNPs and DNA polymerase can significantly influence PCR. First, the addition of DNA polymerase can eliminate the inhibitory effects of excess AuNPs. Second, the addition of AuNPs will increase yield of the desired PCR product and make the optimum concentration of DNA polymerase move to higher value. Third, while excess polymerase might inhibit amplification efficiency, AuNPs can reverse this process and the yield of PCR amplification. Based on these results we propose a possible mechanism that AuNPs might modulate the activity of polymerase and improve PCR amplification.

Nanoparticles Affect PCR Primarily via Surface Interactions with PCR Components: Using Amino-Modified Silica-Coated Magnetic Nanoparticles as a Main Model

“…Similarly, AuNPs, which have been widely reported to enhance the specificity of PCR 10,20,26 , also showed the same manner ( Figure 5D). …”

“…To explore more details about the inhibitory effect on PCR caused by the surface of ASMNPs, we used non-amino-modified nanoparticles as controls. A comparison of Figure 2A and Figure 2B shows that PCR was more sensitive to inhibition caused by ASMNPs than by was previously only attributed to the interaction of the nanomaterials with thermostable DNA polymerase, 26 and other components of the PCR assay mixture were not examined. In the current study, ASMNPs, which are super-paramagnetic, were used to easily identify the components of PCR that were adsorbed by nanomaterials.…”

“…Previously, the effects of nanomaterials on PCR, positive or negative, were all attributed to the binding between nanomaterials and DNA polymerase. [25][26][27] Although our previous results also suggested that ASMNPs could adsorb proteins, such as hydrolyzed skim milk, BSA, and even Taq DNA polymerase, the ability of the binding between ASMNPs and DNA polymerase to positively affect PCR, such as enhancing its specificity, was unclear.…”

“…The main speculations include the following: (1) the effects were the results of binding between the nanomaterial and the DNA polymerase; [25][26][27] (2) 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 5 the function of nanomaterials was similar to that of SSB, which selectively bind single-stranded DNA (ssDNA) and minimize mispairing between primers and templates; 28 and (3) nanomaterials improved the thermal transfer efficiency in the aqueous solution. 13,19 However, these conclusions can not completely explain all of the effects of nanomaterials on PCR.…”

Influences of gold and silver nanoparticles in loop-mediated isothermal amplification reactions

“…As for the non-monotonic behaviour shown in Figure 4, the reduced improvement induced by AuNPs at higher concentration was probably caused by the over-binding of single-stranded of DNA by AuNPs, which leads to the agglomeration and precipitation of AuNPs [21,22]. Another study of normal PCR revealed that the concentrations of both AuNPs and polymerase influence the PCR reaction rate until fully saturated [23].…”