Efficient preparation and properties of mRNAs containing a fluorescent cap analog: Anthraniloyl-m7GpppG

Abstract

AttachedA method has been developed for synthesising fiuorescently labeled capped mRNA. The method incorporates a I single fluorescent molecule as parit of the 5'-mRNA or oligonucleotide cap site. The fluorescent m olecule, Ant-m-GTP Is specifically Incorporated Into the cap site to yield Ant-m7GpppG-capped mRNA or oligonucleotide. Efficient capping f ! was observed with 60^100% of the RNA transcripts capped with the fluorescent m olecule. The Ant-m7G derivative, w hichhas been previously:shown to interact with the eukaryotic cap binding protein elF4E, is shown in this p ap e rto b e : ■ a substrate for the Vaccinia capping enzym e and the DCP2 decapping enzym e from Arabfdopsfs. Further, the Ant- > m7GTP-capped RNA is readily translated. This Ant-m7GTP-capped RNA provides an important tool for monitoring capping reactions, translation, and biophysical studies.