What are the basic principles of real-time PCR?

In real-time PCR (qPCR), the same amplification procedure is used as in conventional PCR. A segment of target DNA, which serves as a template, is combined in a single tube along with other components essential to the amplification reaction (e.g., thermostable DNA polymerases, forward and reverse primers, deoxynucleotide triphosphates (dNTPs), and reaction buffer). The prepared tube is placed in an instrument, referred to as a thermal cycler. A thermal cycler is a laboratory apparatus that typically has a thermal block with holes where tubes holding the PCR reaction mixture are inserted. The reaction contents are then subjected to a series of time and temperature-dependent steps - denaturation, primer annealing, and extension. This series of steps is repeated 25 to 30 times, resulting in an exponential amplification of the DNA template.