Translation of 2′-modified mRNA in vitro and in vivo

2′-Fluoro- and 2′-amino-2′-deoxynucleoside triphosphates have been used for in vitro transcription of 2′-modified luciferase mRNA. The 2′-modified deoxynucleoside-containing transcripts were tested for the expression of luciferase in X.Laevis oocytes as well as in rabbit reticulocyte lysate. Only 2′...

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Bibliographic Details
Published in:Nucleic acids research 1994-11, Vol.22 (23), p.4963-4968
Main Authors: Aurup, Helle, Siebert, Angela, Benseler, Fritz, Williams, David, Eckstein, Fritz
Format: Article
Language:English
Subjects:
Animals
Base Sequence
Cell-Free System
Deoxyribonucleotides - chemical synthesis
Gene Expression Regulation
Luciferases - biosynthesis
Luciferases - genetics
Methionine - metabolism
Molecular Sequence Data
Oocytes
Protein Biosynthesis - physiology
Rabbits
RNA Caps - biosynthesis
RNA, Messenger - chemistry
RNA, Messenger - genetics
Transcription, Genetic
Xenopus laevis
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Summary:2′-Fluoro- and 2′-amino-2′-deoxynucleoside triphosphates have been used for in vitro transcription of 2′-modified luciferase mRNA. The 2′-modified deoxynucleoside-containing transcripts were tested for the expression of luciferase in X.Laevis oocytes as well as in rabbit reticulocyte lysate. Only 2′-fluoro-2′-deoxyadenosine-modified mRNA gave rise to luciferase as shown by SDS gel as well as by enzyme activity measurements in vivo as well as in vitro. 2′-Fluoro-2′-deoxy-pyrimidine nucleoside-modified mRNA did not give rise to luciferase activity. However, they directed incorporation of 35S-labeled methionine into peptide fragments in rabbit reticulocyte lysate indicating premature termination of translation. No or only extremely little of such incorporation could be detected with 2′-amino modified transcripts.
ISSN:0305-1048
1362-4962
DOI:10.1093/nar/22.23.4963