NONHOMOLOGOUS RNA RECOMBINATION IN A CELL-FREE SYSTEM - EVIDENCE FOR A TRANSESTERIFICATION MECHANISM GUIDED BY SECONDARY STRUCTURE

Extensive nonhomologous recombinations occur between the 5' and 3' fra gments of a replicable RNA in a cell-free system composed of pure Q be ta phage replicase and ribonucleoside triphosphates, providing direct evidence for the ability of RNAs to recombine without DNA intermediate s and in the absence of host cell proteins. The recombination events a re revealed by the molecular colony technique that allows single RNA m olecules to be cloned in vitro. The observed nonhomologous recombinati ons are entirely dependent on the 3' hydroxyl group of the 5' fragment , and are due to a splicing-like reaction in which RNA secondary struc ture guides the attack of this 3' hydroxyl on phosphoester bonds withi n the 3' fragment.