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Synthetic mRNAs with Superior Translation and Stability Properties

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Book cover Synthetic Messenger RNA and Cell Metabolism Modulation

Part of the book series: Methods in Molecular Biology ((MIMB,volume 969))

Abstract

The translational efficiency and stability of synthetic mRNA in both cultured cells and whole animals can be improved by incorporation of modified cap structures at the 5′-end. mRNAs are synthesized in vitro by a phage RNA polymerase transcribing a plasmid containing the mRNA sequence in the presence of all four NTPs plus a cap dinucleotide. Modifications in the cap dinucleotide at the 2′- or 3′-positions of m7Guo, or modifications in the polyphosphate chain, can improve both translational efficiency and stability of the mRNA, thereby increasing the amount and duration of protein expression. In the context of RNA-based immunotherapy, the latter is especially important for antigen production and presentation by dendritic cells. Protocols are presented for synthesis of modified mRNAs, their introduction into cells and whole animals, and measurement of their translational efficiency and stability.

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Acknowledgments

This research was supported by grant GM20818 from the National Institutes of Health of the USA (to R.E.R), by the Combined Project Grant SFB 432 from the German Research Foundation (to U.S.), by GO-Bio funding from the German Federal Ministry of Education and Research (to U.S.), by grants NN204 089438 (to J.J.) and NN301 096339 (to E.D.) from the Polish Ministry of Science and Higher Education, and by grant 55005 604 from the Howard Hughes Medical Institute (to E.D.).

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, Lousisiana State University Health Sciences Center, Shreveport, LA, USA

    Ewa Grudzien-Nogalska, Wei Su, Sergey V. Slepenkov & Robert E. Rhoads

  2. Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Warsaw, Poland

    Joanna Kowalska, Edward Darzynkiewicz & Jacek Jemielity

  3. TRON-Translational Oncology at the University Medical Center Mainz, Mainz, Germany

    Andreas N. Kuhn & Ugur Sahin

  4. Ribological, Biontech AG, Mainz, Germany

    Andreas N. Kuhn & Ugur Sahin

  5. III. Medical Department, University Medical Center of the Johannes Gutenberg-University Mainz, Mainz, Germany

    Ugur Sahin

Authors
  1. Ewa Grudzien-Nogalska
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  2. Joanna Kowalska
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  3. Wei Su
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  4. Andreas N. Kuhn
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  5. Sergey V. Slepenkov
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  6. Edward Darzynkiewicz
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  7. Ugur Sahin
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  8. Jacek Jemielity
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  9. Robert E. Rhoads
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Corresponding author

Correspondence to Robert E. Rhoads .

Editor information

Editors and Affiliations

  1. The Anlyan Center, Department of Genetics, Yale School of Medicine, Cedar Street 300, New Haven, 06520-8005, Connecticut, USA

    Peter M. Rabinovich

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Grudzien-Nogalska, E. et al. (2013). Synthetic mRNAs with Superior Translation and Stability Properties. In: Rabinovich, P. (eds) Synthetic Messenger RNA and Cell Metabolism Modulation. Methods in Molecular Biology, vol 969. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-260-5_4

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  • DOI: https://doi.org/10.1007/978-1-62703-260-5_4

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-259-9

  • Online ISBN: 978-1-62703-260-5

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