Protocol

Analysis of RNA by Primer Extension

  1. Joseph Sambrook

Abstract

For mapping the 5′ termini of mRNA molecules, primer extension is the method of choice. A purified oligonucleotide is end-labeled using polynucleotide kinase. The probe and a population of mRNA are allowed to hybridize, and the primers and template are used to carry out reverse transcription using an enzyme cloned from the Moloney murine leukemia virus. The primer extension products are separated on a denaturing polyacrylamide gel and analyzed by radiography.

Footnotes

  • From the Molecular Cloning collection, edited by Michael R. Green and Joseph Sambrook.

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  1. doi:10.1101/pdb.prot101840 Cold Spring Harb Protoc 2021: pdb.prot101840- © 2021 Cold Spring Harbor Laboratory Press
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