käänteiskopiointiPCR

käänteiskopiointiPCR, commonly abbreviated as RT‑PCR, is a laboratory technique used to amplify specific RNA sequences. The method combines reverse transcription, which converts RNA into complementary DNA (cDNA), with the polymerase chain reaction, which amplifies the cDNA to detectable levels. The reverse transcription step is carried out by reverse transcriptase, a ribozyme that reads the RNA template and synthesizes a DNA strand. Once the cDNA is formed, PCR amplification occurs using DNA polymerase, primers that match the target sequence, nucleotides, and a thermocycle that alternates between denaturation, annealing, and extension steps.

The technique is widely employed in molecular biology, diagnostics, and virology. It enables the detection and quantification of viruses such as influenza, HIV, and SARS‑CoV‑2 by measuring viral RNA load. Quantitative RT‑PCR (qRT‑PCR) further incorporates fluorescent dyes or probes, allowing real‑time monitoring of the amplification process and precise determination of initial template amounts.

RT‑PCR differentiation from conventional PCR is essential when the template is RNA rather than DNA. The procedure incorporates additional reagents and time for reverse transcription, but it provides greater sensitivity for low‑copy RNA targets. Since its development in the early 1990s, improvements in reverse transcriptases and probe chemistries have expanded its applications to gene expression profiling, pathogen detection, and biomarker discovery.