EMERGENCE OF A REPLICATING SPECIES FROM AN IN-VITRO RNA EVOLUTION REACTION

The technique of self-sustained sequence replication allows isothermal amplification of DNA and RNA molecules in vitro. This method relief o n the activities of a reverse transcriptase and a DNA-dependent RNA po lymerase to amplify specific nucleic acid sequences. We have modified this protocol to allow selective amplification of RNAs that catalyze a particular chemical reaction. During an in vitro RNA evolution experi ment employing this modified system, a unique class of ''selfish'' RNA s emerged and replicated to the exclusion of the intended RNAs. Member s of this class of selfish molecules, termed RNA Z, amplify efficientl y despite their inability to catalyze the target chemical reaction. Th eir amplification requires the action of both reverse transcriptase an d RNA polymerase and involves the synthesis of both DNA and RNA replic ation intermediates. The proposed amplification mechanism for RNA Z in volves the formation of a DNA hairpin that functions as a template for transcription by RNA polymerase, This arrangement links the two stran ds of the DNA, resulting in the production of RNA transcripts that con tain an embedded RNA polymerase promoter sequence.