Functional messenger RNAs are produced by SP6 in vitro transcription of cloned cDNAs
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We describe a method for the synthesis of microgram quantities of eucaryotic messenger RNAs. Injection into the cytoplasm of frog oocytes and addition to wheat germ extracts show that these synthetic RNAs function efficiently as messenger RNAs. We confirm that a 5' cap on the mRNA is essential for translation in injected oocytes and show that most of the 3' flanking region, including the poly A tail, can be deleted without the abolition of protein synthesis. The method of mRNA synthesis involves in vitro transcription of cDNAs which have been cloned into SP6 vectors (described in the accompanying paper). This method enables one to produce large amounts of mRNA and consequently protein from any cDNA clone.
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First published in 1984, in vitro mRNA transcription (IVT) made transcription of a selected functional mRNA from a template in the desired quantity possible.
“First published in 1984, IVT made the transcription of a selected functional mRNA from a template in the desired quantity possible.20,21”
From COVID-19 mRNA vaccines: Platforms and current developments (2022) · cites this paper as [21]
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- Authors
- Krieg, P. A., & Melton, D. A.
- Affiliations
- Harvard University (US)
- ORCID
- Melton, D.A.
- Venue
- Nucleic Acids Research
- Publisher
- Oxford University Press (OUP)
- Published
- 1984
- Volume
- 12
- Issue
- 18
- Pages
- 7057-7070
- ISSN
- 0305-1048 (ISSN-L)
- 1362-4962
- PMC
- PMC320142
- OpenAlex
- W2064864270
- Cited by
- 1,533
- References
- 32
- Citations by year
- 2026: 10 · 2025: 18 · 2024: 28 · 2023: 15 · 2022: 15 · 2021: 14 · 2020: 6 · 2019: 9 · 2018: 7 · 2017: 2 · 2016: 9 · 2015: 7 · 2014: 7 · 2013: 10 · 2012: 9
- FWCI
- 22.93
- Type
- journal article
- Language
- en
- Open access
- green
- europepmc.org
- Locations
- Nucleic Acids Research (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Research and Splicing
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Molecular Biology Techniques and Applications
- Keywords
- in vitro transcription, SP6 RNA polymerase, cDNA cloning, mRNA 5' cap, polyadenylation, 3' flanking region, mRNA translation, wheat germ extract
- MeSH terms
- Animals, Cloning, Molecular, DNA, Female, Globins, Interferons, Oocytes, Plants, RNA, Messenger, Salmonella Phages, Salmonella typhimurium, Transcription, Genetic, Protein Biosynthesis, Triticum, Xenopus
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