Abstract
The T7 RNA polymerase-dependent transcription was studied as a function of nucleotide sequence structures positioned upstream of the T7 promoter. Model double-stranded DNA templates were constructed for this purpose. They contained a target sequence of 485 base pairs (cDNA fragment of Venesuelian encephalomyelitis equine virus genome), T7 promoter consensus and different extra base sequences upstream of the T7 promoter. The level of the target sequence transcription was clearly determined by the extra base sequence. The presence of one extra base pair G.C ensured the most pronounced effect, transcription was increased one order of magnitude in comparison with template which has only a canonical T7 promoter sequence at the 5'-end.
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- Baranov V. I., Spirin A. S. Gene expression in cell-free system on preparative scale. Methods Enzymol. 1993;217:123–142. doi: 10.1016/0076-6879(93)17059-e. [DOI] [PubMed] [Google Scholar]
- Dickson R. C., Abelson J., Barnes W. M., Reznikoff W. S. Genetic regulation: the Lac control region. Science. 1975 Jan 10;187(4171):27–35. doi: 10.1126/science.1088926. [DOI] [PubMed] [Google Scholar]
- Dunn J. J., Studier F. W. Complete nucleotide sequence of bacteriophage T7 DNA and the locations of T7 genetic elements. J Mol Biol. 1983 Jun 5;166(4):477–535. doi: 10.1016/s0022-2836(83)80282-4. [DOI] [PubMed] [Google Scholar]
- Fahy E., Kwoh D. Y., Gingeras T. R. Self-sustained sequence replication (3SR): an isothermal transcription-based amplification system alternative to PCR. PCR Methods Appl. 1991 Aug;1(1):25–33. doi: 10.1101/gr.1.1.25. [DOI] [PubMed] [Google Scholar]
- Kinney R. M., Johnson B. J., Welch J. B., Tsuchiya K. R., Trent D. W. The full-length nucleotide sequences of the virulent Trinidad donkey strain of Venezuelan equine encephalitis virus and its attenuated vaccine derivative, strain TC-83. Virology. 1989 May;170(1):19–30. doi: 10.1016/0042-6822(89)90347-4. [DOI] [PubMed] [Google Scholar]
- Martin C. T., Coleman J. E. Kinetic analysis of T7 RNA polymerase-promoter interactions with small synthetic promoters. Biochemistry. 1987 May 19;26(10):2690–2696. doi: 10.1021/bi00384a006. [DOI] [PubMed] [Google Scholar]
- Milligan J. F., Groebe D. R., Witherell G. W., Uhlenbeck O. C. Oligoribonucleotide synthesis using T7 RNA polymerase and synthetic DNA templates. Nucleic Acids Res. 1987 Nov 11;15(21):8783–8798. doi: 10.1093/nar/15.21.8783. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rychlik W., Rhoads R. E. A computer program for choosing optimal oligonucleotides for filter hybridization, sequencing and in vitro amplification of DNA. Nucleic Acids Res. 1989 Nov 11;17(21):8543–8551. doi: 10.1093/nar/17.21.8543. [DOI] [PMC free article] [PubMed] [Google Scholar]