Dr. Timofey Evgen'evich Pylaev, IBPPM RAS, Saratov, Russia
Dr. Vladimir Aleksandrovich Bogatyrev, IBPPM RAS, Suratov, Russia
Prof. Nikolai G. Khlebtsov, Institute of Biochemistry and Physiology of Plants and Microorganisms; Saratov State University, Professor, Head of Laboratory of Nanoscale Biosensors (IBPPM), Head of Biophysics Chair (SSU), Saratov, Russia.
Polymerase chain reaction (PCR) is one of the most powerful tools in molecular diagnostics applications. It is widespread in clinical and scientific researches for sensitive and rapid DNA detection. Typical conventional PCR assay consists of two stages: amplification and registration of the amplified PCR-products (amplicons). The last stage can be performed by the agarose gel-electrophoresis with ethidium bromide staining. The main problem of conventional PCR is low specificity in non-optimized systems that results in broad smearing bands through the whole agarose lane. Great efforts were done to enhance the specificity, and a wide list of additives for PCR-mixture was suggested. These additives include small organic molecules (DMSO, glycerol, formamide), detergents (Triton X-100, Tween-20), and proteins (bovine serum albumin (BSA), single-stranded DNA binding protein (SSB)) . Recently, a novel class of PCR enhancers such as metal nanoparticles (NPs), mainly AuNPs , was introduced in the error-prone PCR. One of the problems unsolved to the date are the mechanisms underlying these processes. The speculative mechanisms are related to the interaction between the components of PCR mixture and the AuNPs, the changes in polymerase activity by the AuNPs, the SSB-like mechanism, and the heat transfer properties of AuNPs . The main goal of the present study was to improve the non-specific PCR systems and to explore the key mechanisms for underlying processes in the amplification and detection stages of PCR.
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Dr. Timofey Evgen'evich Pylaev
IBPPM RAS, PhD, research assistant in Lab of Nanobiotechnology, IBPPM RAS
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