Introduction
More than 15 years after its initial description, the polymerase chain reaction (PCR) has become a standard molecular
biology tool and is the most widely used method of nucleic acid amplification. The procedure is based on the
ability of DNA polymerase to copy a strand of DNA by elongation of a complementary oligonucleotide primer. As
use of PCR has increased, scientists have improved upon the initial PCR method with valuable modifications. Today
PCR is performed in multiple formats including single tubes and multi-well plates. Fairly precise quantitative techniques
have been developed from the basic technology allowing the amplification process to be monitored. The PCR technique
has become an important basis for an expanding number of molecular diagnostics applications.
The Basics
As originally described, PCR exploits the DNA replication mechanisms of the cell. Using a thermostable DNA polymerase,
two oligonucleotide primers (usually between 18 and 28 nucleotides in length) and deoxynucleotide triphosphates
(dNTPs), a specific DNA sequence is amplified exponentially. The primers flank the region of interest and are
complementary to opposite strands of the template DNA (see Figure 1). PCR involves repeated replication of template
DNA via a series of cycles. Each cycle consists of three stages:
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