Preparation of Functional, Fluorescently Labeled mRNA Capped with Anthraniloyl-m(7)GpppG

Methods Mol Biol. 2016:1428:61-75. doi: 10.1007/978-1-4939-3625-0_4.

Abstract

Fluorescent mRNA molecules offer a wide range of applications for studying capping/decapping reactions, translation, and other biophysical studies. Furthermore, fluorescent tags prove invaluable for tracking RNA molecules in cells. Here, we describe an efficient synthesis of a fluorescent cap analog, anthranioyl-GTP, its purification, and in vitro cap labeling of transcribed mRNA catalyzed by the recombinant vaccinia capping enzyme to produce anthranioyl-m(7)GpppG-capped RNA.

Keywords: Ant-GTP; Anthranioyl-m7G; Capped RNA; Chromatography; Fluorescence; Fluorescently labeled RNA; RNA labeling.

MeSH terms

  • Guanosine / analogs & derivatives
  • Guanosine / chemistry
  • Molecular Structure
  • Protein Biosynthesis
  • RNA Cap Analogs / chemical synthesis*
  • RNA Cap Analogs / chemistry
  • RNA, Messenger / chemistry*
  • RNA, Messenger / genetics
  • Spectrometry, Fluorescence
  • Transcription, Genetic

Substances

  • RNA Cap Analogs
  • RNA, Messenger
  • Guanosine
  • 7-methylguanosine