Template-free RNA synthesis by Qβ replicase

Abstract

In the absence of extraneously added template, standard preparations of Qβ replicase spontaneously synthesize RNA in vitro, possibly as a result of RNA contamination¹. Using special enzyme purifications, Sumper and Luce² 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^3–5, the products synthesized ‘de novo’ by Qβ 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 parameters⁶. Kinetic analysis of the de novo processes⁷ distinguished it from template-instructed synthesis and excluded an assumption of self-replicating RNA contamination. These conclusions were questioned recently by Hill and Blumenthal⁸, who claimed to show that highly purified Qβ replicase preparations cannot produce RNA de novo. We now present evidence that, under the conditions required for de novo synthesis, Qβ replicase prepared according to their method is also capable of de novo synthesis. Furthermore, we show that Qβ replicase condenses nucleoside triphosphates to more or less random oligonucleotides.