Polymerase chain reaction

PCR is a new achievement in molecular genetics, used for DNA amplification and allows the rapid expansion of the specific

DNA region( ie any gene of interest) in vitro more than 200,000 times. To carry out the reaction, it is sufficient to have the DNA material of one cell;the amount of DNA amplified by PCR is so great that this DNA can simply be stained( using radioactive probes after electrophoresis is not required).A prerequisite for conducting PCR is the knowledge of the nucleotide sequence of the amplified DNA region for the correct selection of artificially synthesized primers.

Currently, PCR is a process that takes place in a single tube and consists of repeated cycles of amplification( duplication, copying) of the specific sequence of the DNA molecule in order to obtain a sufficiently large number of copies that can be identified by electrophoresis. One of the key components of the reaction is "primers" - synthetic oligonucleotides, consisting of 20-30 bases, complementary to "sites"( sections) of annealing( attachment) at the identified site of the template DNA.

PCR runs automatically in a programmable thermostat - thermocycler( thermocycler).The three-step cycle, which results in replicas of the identifiable portion of the template DNA, is repeated 30-50 times in accordance with the preset program of the thermal cycler. In the first cycle, the oligopramers hybridize with the original template DNA, and then( in subsequent cycles) and with the newly synthesized DNA molecules as they accumulate in the reaction mixture. In the latter case, the synthesis of DNA ends not due to a change in the temperature regime, but upon reaching the DNA polymerase boundary of the amplified region, which determines the size of the newly synthesized DNA region to within one nucleotide.

As a method of detecting the obtained DNA molecules, electrophoresis is used, by means of which the amplified material is divided according to the size of amplicons( amplification products).

With the help of PCR, it is possible to directly investigate the sites of localization of suspected mutations or polymorphic sites, and also to study the presence of any other specific features of DNA.