reversetranscription

Reverse transcription is the process of synthesizing complementary DNA (cDNA) from an RNA template, performed by reverse transcriptase enzymes. It is a key step in converting RNA into a DNA form that can be amplified, cloned, or sequenced. This is distinct from transcription, where DNA is used to make RNA.

Enzymes and contexts. Reverse transcriptases are found in retroviruses, retroelements, and specialized cellular enzymes such as telomerase. Common laboratory enzymes include MMLV reverse transcriptase and avian myeloblastosis virus (AMV) reverse transcriptase, as well as engineered thermostable variants. Telomerase is a dedicated reverse transcriptase that extends chromosome ends using an RNA template inside the enzyme complex.

Typical workflow. In the lab, RNA is primed with oligo-dT, random hexamers, or gene-specific primers, and reverse transcriptase synthesizes a single-stranded cDNA. Often a second-strand synthesis step converts this into double-stranded DNA suitable for PCR, cloning, or sequencing. The quality of the RNA, primer choice, and enzyme fidelity influence the accuracy and completeness of the resulting cDNA.

Applications and considerations. Reverse transcription is essential for analyzing gene expression (via RT-PCR and RT-qPCR), RNA sequencing library preparation, and diagnostics for RNA viruses. Enzymes vary in fidelity and have RNase H activity; some high-fidelity or thermostable variants reduce errors but may require optimized conditions. In biology, reverse transcription also underpins telomere maintenance through the action of telomerase.