Template-free RNA synthesis by Q beta replicase

Abstract

In the absence of extraneously added template, standard preparations of Q beta replicase spontaneously synthesize RNA in vitro, possibly as a result of RNA contamination. Using special enzyme purifications, Sumper and Luce presented evidence that self-replicating RNA not present ab initio can grow out of 'template-free' incorporation mixtures. In contrast to DNA polymerase I and RNA polymerase, which also show de novo synthesis, the products synthesized 'de novo' by Q beta replicase are RNA species containing nonrepetitive sequences of defined lengths which differ between experiments, even when synthesized under identical conditions, in fingerprints, chain lengths and kinetic parameters. Kinetic analysis of the de novo processes distinguished it from template-instructed synthesis and excluded an assumption of self-replicating RNA contamination. These conclusions were questioned recently by Hill and Blumenthal, who claimed to show that highly purified Q beta replicase preparations cannot produce RNA de novo. We now present evidence that, under the conditions required for de novo synthesis, Q beta replicase prepared according to their method is also capable of de novo synthesis. Furthermore, we show that Q beta replicase condenses nucleoside triphosphates to more or less random oligonucleotides.