A method that uses the enzyme Reverse Transcriptase (RNA-dependent DNA polymerase) to determine the 5′-untranslated region of specific mRNA molecule
Primer extension is a laboratory technique that makes use of the enzyme reverse transcriptase (or the RNA-dependent DNA polymerase). It was discovered by Howard Temin and David Baltimore in 1970. It is used for measuring the amount of RNA and for mapping the 5’ends of RNA. It is also used to create cDNA libraries from mRNA. It is also for the detection of transcripts produced during in vitro transcription and for mapping the replication initiation point.1
The primer extension is an alternative method to nuclease protection assay (e.g. S1 nuclease mapping) when wanting to quantify and map RNA transcripts. Both of them are capable of determining where mRNA starts and an estimate of the concentration of a transcript based on the intensity of the transcript band from autoradiograph.
The advantage of primer extension method is that there is no probe used and it is simple and straight forward. The drawback is the reverse transcriptase that may encounter regions of RNA that can pause or terminate synthesis. 1
1Alice Mumbey-Wafula. 20 February 2004. Primer extension, and nuclease si mapping. – Biology. Biochemistry Seminar.