Polymerase Chain Reaction

organism number to readily detectable levels, yet culture is not always easy or successful. A novel technique, the polymerase chain reaction (PCR), was recently developed for in vitro amplification of the DNA or RNA of an organism or gene defect, and culture may not be required. The PCR takes advantage of an enzyme that uses a defined segment in a strand of DNA as a template for assembling a complementary strand. The principle of the PCR is simple, requiring a three-step cycling process: (7) denaturation of doublestranded DNA, (2) annealing of primers, and (3) primer extension. If an RNA sequence is to be amplified, a DNA copy of it (cDNA) must be synthesized by using reverse transcriptase before the PCR is begun. A cycle typically takes ~3-5 min and is repeated 20-40 times 1. The PCR reaction vessel contains a mixture of buffers, nucleotides, primers, enzyme, and nucleic acid from the specimen of interest.