Polymerase chain reaction (PCR) is a useful molecular biology technique developed by Kary Mullis in the 1980s. PCR is used to amplify a particular DNA sequence to produce millions of copies within hours.
Due to its powerful ability, PCR is applied in different areas such as; genetic and infectious disease identification, forensics, and research in molecular biology. In contrast to conventional PCR, quantitative reverse transcriptase PCR (qRT-PCR) allows the quantitative measurement of gene expression in real-time.
In general, a complete PCR reaction requires five basic PCR reagents; a DNA/RNA template, DNA polymerase, primers (forward and reverse), deoxynucleotide triphosphates (dNTPs) and PCR buffers.
Selecting the right templates for PCR is vital, as different templates are required depending on the method of PCR. For instance, when performing RT-qPCR, RNA templates are required for producing complementary DNA whereas DNA templates are needed for conventional PCR. It is also important to have high quality DNA/RNA templates in large quantities to help optimize the efficiency of PCR. For this reason, a good PCR template preparation kit is needed for the first step of a successful PCR.
The initiation of DNA synthesis requires primers; short strands of nucleotides (DNA or RNA) which are complementary to the template DNA and serve as a DNA synthesis starting point for the DNA/RNA polymerase. Annealing primers to single strand DNA requires lower temperature (50-65 °C) than the denaturation step. Once the annealing step is completed, hydrogen bonds will form between the primers and the template DNA.
