Superior 5' homogeneity of RNA from ATP-initiated transcription under the T7 2.5 promoter
- 2 January 2004
- journal article
- Published by Oxford University Press (OUP) in Nucleic Acids Research
- Vol. 32 (1), 14e-14
- https://doi.org/10.1093/nar/gnh007
Abstract
Transcription from the commonly used GTP- initiating T7 class III promoter phi6.5 frequently produces heterogeneous RNA at both 3' and 5' ends. We demonstrate here that RNA transcripts from the T7 class II promoter phi2.5 have superior 5' homogeneity over those from the phi6.5 promoter, with comparable total RNA yields. The overall homogeneity of RNA transcripts is improved to different degrees depending on RNA sequences, although transcription under phi2.5 does not affect the 3' heterogeneity of RNA. In combination with 3' RNA trimming by DNAzymes or ribozymes, this ATP- initiated transcription system based on the T7 phi2.5 promoter can provide excellent quality of RNA for applications requiring a high degree of RNA size homogeneity.Keywords
This publication has 26 references indexed in Scilit:
- Synthesis of adenosine derivatives as transcription initiators and preparation of 5′ fluorescein- and biotin-labeled RNA through one-step in vitro transcriptionRNA, 2003
- NMR Spectroscopy of RNAChemBioChem, 2003
- A Simple and Efficient Method to Transcribe RNAs with Reduced 3′ HeterogeneityMethods, 2001
- RNA-Catalyzed CoA, NAD, and FAD Synthesis from Phosphopantetheine, NMN, and FMNBiochemistry, 2000
- A Preorganized Active Site in the Crystal Structure of the Tetrahymena RibozymeScience, 1998
- Crystallization of RNA-protein complexes I. Methods for the large-scale preparation of RNA suitable for crystallographic studiesJournal of Molecular Biology, 1995
- [13] Preparation of isotopically enriched RNAs for heteronuclear NMRMethods in enzymology, 1995
- Three-dimensional structure of a hammerhead ribozymeNature, 1994
- Three-dimensional heteronuclear NMR studies of RNANature, 1992
- Complete nucleotide sequence of bacteriophage T7 DNA and the locations of T7 genetic elementsJournal of Molecular Biology, 1983