RNA Template-Directed RNA Synthesis by T7 RNA Polymerase

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 91; no. 15; pp. 6972 - 6976
• Main Authors: Cazenave, Christian, Uhlenbeck, Olke C.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences of the United States of America 19.07.1994; National Acad Sciences; National Academy of Sciences
• Subjects: Bacteria; Base Sequence; Biochemistry; DNA; DNA-Directed RNA Polymerases - metabolism; Enzymes; Gels; Molecular Sequence Data; Molecules; Nucleic acids; Nucleotides; Oligomers; Oligonucleotides; phage T7; Ribonucleic acid; RNA; RNA - biosynthesis; Templates, Genetic; Transcription, Genetic; Viral Proteins
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.91.15.6972

In an attempt to synthesize an oligoribonucleotide by run-off transcription by bacteriophage T7 RNA polymerase, a major transcript was produced that was much longer than expected. Analysis of the reaction indicated that the product resulted from initial DNA-directed run-off transcription followed by RNA template-directed RNA synthesis. This reaction occurred because the RNA made from the DNA template displayed self-complementarity at its 3' end and therefore could form an intra- or intermolecular primed template. In reactions containing only an RNA template, the rate of incorporation of NTPs was quite comparable to DNA-dependent transcription. RNA template-directed RNA synthesis has been found to occur with a great number of oligoribonucleotides, even with primed templates that are only marginally stable. In one instance, we observed a multistep extension reaction converting the oligonucleotide into a final product longer than twice its original length. Presumably, such a process could have generated some of the RNAs found to be efficiently replicated by T7 RNA polymerase.