PCR Amplification and Transcription for Site-Specific Labeling of Large RNA Molecules by a Two-Unnatural-Base-Pair System
Joint Authors
Hirao, Ichiro
Yokoyama, Shigeyuki
Yamashige, Rie
Kimoto, Michiko
Source
Issue
Vol. 2012, Issue 2012 (31 Dec. 2012), pp.1-8, 8 p.
Publisher
Hindawi Publishing Corporation
Publication Date
2012-06-26
Country of Publication
Egypt
No. of Pages
8
Main Subjects
Abstract EN
For the site-specific labeling and modification of RNA by genetic alphabet expansion, we developed a PCR and transcription system using two hydrophobic unnatural base pairs: 7-(2-thienyl)-imidazo[4,5-b]pyridine (Ds) and 2-nitro-4-propynylpyrrole (Px) as a third pair for PCR amplification and Ds and pyrrole-2-carbaldehyde (Pa) for the incorporation of functional components as modified Pa bases into RNA by T7 transcription.
To prepare Ds-containing DNA templates with long chains, the Ds-Px pair was utilized in a fusion PCR method, by which we demonstrated the synthesis of 282-bp DNA templates containing Ds at specific positions.
Using these Ds-containing DNA templates and a biotin-linked Pa substrate (Biotin-PaTP) as a modified Pa base, 260-mer RNA transcripts containing Biotin-Pa at a specific position were generated by T7 RNA polymerase.
This two-unnatural-base-pair system, combining the Ds-Px and Ds-Pa pairs with modified Pa substrates, provides a powerful tool for the site-specific labeling and modification of desired positions in large RNA molecules.
American Psychological Association (APA)
Kimoto, Michiko& Yamashige, Rie& Yokoyama, Shigeyuki& Hirao, Ichiro. 2012. PCR Amplification and Transcription for Site-Specific Labeling of Large RNA Molecules by a Two-Unnatural-Base-Pair System. Journal of Nucleic Acids،Vol. 2012, no. 2012, pp.1-8.
https://search.emarefa.net/detail/BIM-455750
Modern Language Association (MLA)
Kimoto, Michiko…[et al.]. PCR Amplification and Transcription for Site-Specific Labeling of Large RNA Molecules by a Two-Unnatural-Base-Pair System. Journal of Nucleic Acids No. 2012 (2012), pp.1-8.
https://search.emarefa.net/detail/BIM-455750
American Medical Association (AMA)
Kimoto, Michiko& Yamashige, Rie& Yokoyama, Shigeyuki& Hirao, Ichiro. PCR Amplification and Transcription for Site-Specific Labeling of Large RNA Molecules by a Two-Unnatural-Base-Pair System. Journal of Nucleic Acids. 2012. Vol. 2012, no. 2012, pp.1-8.
https://search.emarefa.net/detail/BIM-455750
Data Type
Journal Articles
Language
English
Notes
Includes bibliographical references
Record ID
BIM-455750