Product analysis of RNA generated de novo by Qβ replicase

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Abstract

Qβ replicase synthesizes self-replicating RNA in the absence of exogenous template after a certain lag time (“template-free synthesis”). The products of the template-free RNA synthesis have been investigated by gel electrophoresis and fingerprinting techniques. It has been found that a multitude of self-replicating RNA species appears in the early phases of reaction with variable lengths and sequences. Template-free synthesis in different samples under completely identical conditions yields RNA products with very different and unrelated fingerprints. The early products rapidly undergo an evolution process that alters the phenotypic properties of the self-replicating RNA, and leads to a concomitant increase of replication efficiency. Fingerprints and electrophoretic mobilities of the self-replicating RNA species are altered discontinuously during the evolution process. The evolution process ends with the selection of optimized self-replicating RNA species, whose phenotypes are conserved even after many serial transfers. Some optimized RNA species and midivariant RNA apparently have related sequences, since they contain many identical spots in their fingerprints. The properties of the RNA species produced by template-free synthesis match those of 6 S RNA found in Qβ-infected Escherichia coli cells. The results are in full agreement with the finding of Sumper & Luce (1975), who have presented evidence that Qβ replicase synthesized RNA de novo in the absence of exogenous template.

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