RNA In Vitro Synthesis by Phage T7 DNA-Dependent RNA Polymerase
The purpose of this chapter is to introduce beginners in molecular biology to RNA transcription by phage T7 DNA-dependent RNA polymerase. The work outlined here includes the transcription procedure of plasmid vectors or PCR-amplified DNA templates, the purification and identification of RNA products by sequencing with reverse transcriptase.
KeywordsBacteriophage Promoter Extension Inhibition Transcription Vector Secondary Structure Effect Electrophoretic Concentrator
Unable to display preview. Download preview PDF.
- Hartz D, McPheeters DS, Traut R, Gold L (1988) Extension inhibition analysis of translation initiation complexes. In: Noller HF Jr, Moldave K (eds) Methods Enzymology 164: 419–425Google Scholar
- Howe, CJ, Ward ES (1989) Nucleic Acid Sequencing. Oxford University Press, Oxford, pp 30–75Google Scholar
- Klement JF, Moorefield MB, Jorgensen E, Brown JE, Risman S, McAllister WT (1990) Discrimination between bacteriophage T3 and T7 promoters by the T3 and T7 RNA polymerases depends primarily upon a three base pair region located 10 to 12 base-pairs upstream from the start site. J Mol Biol 215: 21–29PubMedCrossRefGoogle Scholar
- Mackie GA (1988) Vectors for the synthesis of specific RNAs in vitro. In: Rodriguez RL, Denhardt DT (eds) A survey of molecular cloning vectors and their uses. Butterworths, New York, pp 253–267Google Scholar
- Maniatis T, Fritsch EF, Sambroock (1982) Molecular cloning: a laboratory manual. Cold Spring Harbor, New York, pp 1. 25–1. 52Google Scholar