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Updated: Jul 24, 2026

Toeprinting Analysis of Translation Initiation Complex Formation on Mammalian mRNAs
Published on: May 10, 2018
Facile characterization of translation initiation via nonsense codon suppression
A V Karginov1, M Lodder, S M Hecht
1Department of Chemistry and Department of Biology, University of Virginia, Charlottesville, VA 22901, USA.
Abstract:
A new strategy for studying the mechanism of translation initiation in eukaryotes has been developed. The strategy involves the use of an in vitro translation system to incorporate a non-natural fluorescent amino acid into a protein from a suppressor tRNAPheCUA misacylated with that amino acid. It is thereby possible to monitor translation initiation efficiency at an AUG codon in different contexts; this is illustrated for three constructs encoding Escherichia coli dihydrofolate reductase mRNA with different translation initiation regions. Fluorescence measurements after in vitro translation of the mRNAs in rabbit reticulocyte lysate reflected differences in the position and efficiency of translation initiation and, therefore, can be used for characterization of the translation initiation process.
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