In vivo duplication of genetic elements by the formation of stem-loop DNA without an RNA intermediate.

Abstract

Gene duplication through cDNA synthesis by reverse transcriptase is believed to have played an important role in the diversification of genomes during evolution. Here, we demonstrate that a genomic DNA sequence can be duplicated in vivo as a result of template switching. When an inverted repeat (IR) structure was inserted in a site downstream from a ColE1 plasmid origin of DNA replication, transformation of Escherichia coli cells with this plasmid resulted in the production of a new DNA fragment encompassing the region from the origin to the center of the IR structure. The structure of this DNA molecule is composed of a long stem-loop formed by a single-stranded DNA, in which the loop is formed by the IR structure. The DNA fragment is designated slDNA, for stem-loop DNA. The experiments in this study suggest that during DNA replication, template switching at the stem-loop structure formed by the IR structure gives rise to slDNA utilizing the nascent DNA strand or the parental strand as a template. The mechanistic implications of slDNA synthesis, and its possible roles in genome evolution, are discussed.