PMID: 7524080Sep 27, 1994Paper

Coupling of RNA displacement and intrinsic termination in transcription from synthetic RNA.DNA bubble duplex constructs

Proceedings of the National Academy of Sciences of the United States of America
S S DaubePeter H von Hippel

Abstract

Functional transcription elongation complexes can be formed by adding RNA polymerase in trans to a preformed nucleic acid construct. This construct consists of a double-stranded DNA fragment that contains a noncomplementary (permanent DNA bubble) region into which an RNA primer oligonucleotide has been hybridized. By ligating a DNA fragment containing the strong intrinsic terminator T7Te to the RNA.DNA bubble duplex, we show here that Escherichia coli core RNA polymerase-catalyzed transcription, initiated from such a construct, terminates at the predicted position. Furthermore, we show that the termination efficiency obtained is comparable to that observed in a control reaction initiated with the E. coli holopolymerase from the T7A1 promoter if an RNA oligomer trap is used to permit proper displacement of the nascent RNA from the DNA template strand. The trap oligomer is complementary to the template strand of the permanent DNA bubble and prevents rehybridization of the nascent RNA at this site. Varying the amount of RNA trap that is added permits us to modulate the extent of total RNA displacement. Our results show that RNA displacement and termination efficiency are directly correlated, suggesting that intrinsic termination r...Continue Reading

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Citations

Jun 16, 2006·RNA·Maxim V SukhodoletsSankar Adhya
Jun 24, 1997·Proceedings of the National Academy of Sciences of the United States of America·P R Bianco, S C Kowalczykowski
Jul 21, 1999·Proceedings of the National Academy of Sciences of the United States of America·S S Daube, P H von Hippel
Aug 1, 2006·Journal of Molecular Biology·Innokenti Toulokhonov, Robert Landick
Apr 4, 2006·Biophysical Journal·Oana CobanG Ulrich Nienhaus
Jan 20, 2018·Physical Review. E·Panayotis Benetatos
Sep 4, 2007·Cell and Tissue Research·Yoshiki HosodaSeiji Matsuda
Apr 17, 1999·The Journal of Biological Chemistry·R G Keene, D S Luse

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