Biomics

Despite the apparent ease of conducting PCR with a pair of primers, experimenters often encounter the fact that even well-chosen primers do not provide the desired result, which can occur for many reasons. These include the amplification of so-called "difficult" templates, which include those with a high GC composition of the amplified DNA region, which causes the formation (after the denaturation stage) of strong secondary structures in single-stranded DNA, which DNA polymerase is sometimes unable to overcome at the elongation stage, as well as an excessively large length of amplicons with low processivity of enzyme. In addition, DNA polymerase is sometimes inhibited due to certain substances present in the reaction mixture (usually added together with the analyzed DNA preparation) and interfering with the full functioning of this enzyme. To overcome these difficulties, it is proposed to add in the reaction mixture additional components or enhancers, which may include various nanomaterials and nanocomposite substances, which led to the appearance of the so-called nanoPCR. Among them the most popular additives are gold nanoparticles, including those coated with various compounds that change their surface charge. Nanoparticles of some metals or their oxides or sulfides are also widely used in nanoPCR. Various allotropic forms of carbon are also used – graphene, graphene oxide, reduced graphene oxide, diamond nanoparticles, carbon nanotubes, carbon nanopowder. There were attempts to use fullerene, but it only inhibited the reaction. In addition to these nanocompounds, various quantum dots of different diameters and compositions, as well as some other nanoscale substances, are often used. The mechanisms of action in nanoPCR of these nanocompounds differ, primarily due to different structures, but several types can be distinguished in the form of sorption on their surfaces of both single-stranded DNA and DNA polymerase, as well as an increase in the thermal conductivity of the solution. At the same time, only a slight excess of the optimal concentrations of such enhancers can lead to inhibition of the amplification process.

PCR, nanoPCR, enhancer, nanomaterials

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