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Updated: Mar 27, 2026

In vitro Transcription and Capping of Gaussia Luciferase mRNA Followed by HeLa Cell Transfection
Published on: March 26, 2012
Efficient preparation and properties of mRNAs containing a fluorescent cap analog: Anthraniloyl-m(7)GpppG
Dilantha Gunawardana1, Artem V Domashevskiy2, Ken R Gayler3
1Department of Biochemistry and Molecular Biology; Bio21 Molecular Science and Biotechnology Institute; University of Melbourne, Parkville; Victoria, Australia; Present address: Department of Botany; University of Sri Jayewardenepura; Soratha Mawatha, Nugegoda, Sri Lanka.
Abstract:
A method has been developed for synthesising fluorescently labeled capped mRNA. The method incorporates a single fluorescent molecule as part of the 5'-mRNA or oligonucleotide cap site. The fluorescent molecule, Ant-m(7)GTP is specifically incorporated into the cap site to yield Ant-m(7)GpppG-capped mRNA or oligonucleotide. Efficient capping was observed with 60-100% of the RNA transcripts capped with the fluorescent molecule. The Ant-m(7)G derivative, which has been previously shown to interact with the eukaryotic cap binding protein eIF4E, is shown in this paper to be a substrate for the Vaccinia capping enzyme and the DCP2 decapping enzyme from Arabidopsis. Further, the Ant-m(7)GTP-capped RNA is readily translated. This Ant-m(7)GTP-capped RNA provides an important tool for monitoring capping reactions, translation, and biophysical studies.
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