Site-directed mutagenesis: effect of an extracistronic mutation on the in vitro propagation of bacteriophage Qbeta RNA.

It has been proposed that the nucleotide sequences of the 3' terminal extracistronic regions of phage RNA plus and minus strands have been strictly conserved during evolution because they are stringently required for recognition by the viral replicase. We have devised a method to generate point mutations at selected sites in the genome of phage Qbeta. The in vitro synthesis of Qbeta RAN with G leads to A transition in the 16th position from the 3' end, i.e., in the terminal extracistronic region of the genome, is outlined. When a mixture of about 60% wild-type and 40% mutant RNA was repeatedly replicated, the mutant RNA was enriched to 80%, showing that at least this point mutation in the terminal sequence of Qbeta RNA does not impair its in vitro replication, but in fact slightly accelerates it.