Development Application of Polymerase Chain Reaction (PCR)

Polymerase chain reaction (PCR) is an in vitro technique to synthesize large quantities of a given DNA molecule that separates the DNA into two complementary strands, uses a peltier heat pump to quickly heat and cool the DNA and uses the Taq polymerase for the synthesis of DNA. Taq is a bacterium that lives by volcanic sulfer jets at the bottom of the ocean where the temperature is very high. For reverse transcription PCR, primers are short strands of RNA that bind to the target site of DNA molecule. DNA polymerases need to have RNA primers for the beginning of DNA replication. Four dNTPs (deoxyribonucleotide triphosphates) (dGTP, dCTP, dATP and dTTP) are bricks of the DNA molecules and the Taq polymerase uses the dNTPs to build the new DNA molecular chains. The real-time PCR (RT-PCR), also called quantitative RT-PCR (qRT-PCR) or kinetic PCR (kPCR), is a technique used to simultaneously quantify and amplify a DNA molecule, and it is used to determine whether a specific DNA sequence is present in the sample (if it is present, the number of copies in the sample). The procedure of RT-PCR follows the regular PCR procedure, but the DNA is quantified after each round of amplification. Two common methods of quantification are the use of fluorescent dyes that intercalate with double-strand DNA, and modified DNA oligonucleotide probes that fluoresce when hybridized with a complementary DNA. RT-PCR could be combined with reverse transcription PCR to quantify messenger RNA (mRNA) at a particular time for in a particular cell or tissue type. [The Journal of American Science. 2005;1(3):1-47].