ArticleRNA (New York, N.Y.)2016
A general method for rapid and cost-efficient large-scale production of 5' capped RNA.
Article in RNA (New York, N.Y.), 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 58 papers.
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Who cites it
58 citing papers in PubMed, 104 citations in OpenAlex.
- Beyond the genetic code: orchestrating epigenetic and immune landscapes with multivalent mRNA-exosome vaccines.Precision clinical medicine · 2026Review
- Transcript-independent quantitative analysis of RNA cap structures by LC-MS/MS.Molecular therapy. Advances · 2026Article
- Engineered T7 RNA polymerase to improve mRNA capping efficiency and reduce dsRNA generation duringMolecular therapy. Advances · 2026Article
- Beyond the Sequence: Chemical and Topological Design and Innovations in mRNA Therapeutics.Chemical reviews · 2026Review
- "More" Artificial mRNAs: Beyond the Art of Nature.Advanced science (Weinheim, Baden-Wurttemberg, Germany) · 2026Review
- Novel trinucleotide mRNA capping reagents: improved synthetic route and efficient cotranscriptional incorporation in mRNA.RNA (New York, N.Y.) · 2026Article
- The mRNA-Based Innovative Strategy: Progress and Challenges.Nano-micro letters · 2026Review
- RNA vaccines: The dawn of a new age for tuberculosis?Human vaccines & immunotherapeutics · 2025Review
- Article
- Quantitative profiling of human translation initiation reveals elements that potently regulate endogenous and therapeutically modified mRNAs.Molecular cell · 2025Article
- Mechanistic insights into Sindbis virus infection: noncapped genomic RNAs enhance the translation of capped genomic RNAs to promote viral infectivity.Nucleic acids research · 2025Article
- mRNA vaccines in the context of cancer treatment: from concept to application.Journal of translational medicine · 2025Review
- Optimization of In Vitro Transcription Reaction for mRNA Production Using Chromatographic At-Line Monitoring.Methods in molecular biology (Clifton, N.J.) · 2025Article
- Current Non-Viral-Based Strategies to Manufacture CAR-T Cells.International journal of molecular sciences · 2024Review
- mRNA cancer vaccines from bench to bedside: a new era in cancer immunotherapy.Biomarker research · 2024Review
- Personalized mRNA vaccines in glioblastoma therapy: from rational design to clinical trials.Journal of nanobiotechnology · 2024Review
- mRNA medicine: Recent progresses in chemical modification, design, and engineering.Nano research · 2024Article
- Advances in Nonviral mRNA Delivery Materials and Their Application as Vaccines for Melanoma Therapy.ACS applied bio materials · 2024Review
- Role of RNA structural plasticity in modulating HIV-1 genome packaging and translation.Proceedings of the National Academy of Sciences of the United States of America · 2024Article
- Supercoiled DNA percentage: A key in-process control of linear DNA template for mRNA drug substance manufacturing.Molecular therapy. Nucleic acids · 2024Article
Corrections and comments
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
Abstract
The eukaryotic mRNA 5' cap structure is indispensible for pre-mRNA processing, mRNA export, translation initiation, and mRNA stability. Despite this importance, structural and biophysical studies that involve capped RNA are challenging and rare due to the lack of a general method to prepare mRNA in sufficient quantities. Here, we show that the vaccinia capping enzyme can be used to produce capped RNA in the amounts that are required for large-scale structural studies. We have therefore designed an efficient expression and purification protocol for the vaccinia capping enzyme. Using this approach, the reaction scale can be increased in a cost-efficient manner, where the yields of the capped RNA solely depend on the amount of available uncapped RNA target. Using a large number of RNA substrates, we show that the efficiency of the capping reaction is largely independent of the sequence, length, and secondary structure of the RNA, which makes our approach generally applicable. We demonstrate that the capped RNA can be directly used for quantitative biophysical studies, including fluorescence anisotropy and high-resolution NMR spectroscopy. In combination with (13)C-methyl-labeled S-adenosyl methionine, the methyl groups in the RNA can be labeled for methyl TROSY NMR spectroscopy. Finally, we show that our approach can produce both cap-0 and cap-1 RNA in high amounts. In summary, we here introduce a general and straightforward method that opens new means for structural and functional studies of proteins and enzymes in complex with capped RNA.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.