Integration of Bombyx mori R2 sequences into the 28S ribosomal RNA genes of Drosophila melanogaster.
R2 non-long-terminal-repeat retrotransposable elements integrate into a precise location in the 28S rRNA genes of arthropods. The purified protein encoded by R2 can cleave the 28S gene target site and use the 3' hydroxyl group generated by this cleavage to prime reverse transcription of its own RNA, a process called target-primed reverse transcription. An integration system is described here in which components from the R2 element of the silkmoth, Bombyx mori, are injected into the preblastoderm embryo of Drosophila melanogaster. Silkmoth R2 sequences were readily detected in the 28S rRNA genes of the surviving adults as well as in the genes of their progeny. The 3' junctions of these insertions were similar to those seen in our in vitro assays, as well as those from endogenous R2 retrotransposition events. The 5' junctions of the insertions originally contained major deletions of both R2 and 28S gene sequences, a problem overcome by the inclusion of upstream 28S gene sequences at the 5' end of the injected RNA. The resulting 5' junctions suggested a recombination event between the cDNA and the upstream target sequences. This in vivo integration system should help determine the mechanism of R2 retrotransposition and be useful as a delivery system to integrate defined DNA sequences into the rRNA genes of organisms.
Eickbush
D G
DG
Department of Biology, University of Rochester, Rochester, New York 14627-0211, USA.
Luan
D D
DD
Eickbush
T H
TH
D000818:Animals; D030801:Animals, Genetically Modified; D012831:Bombyx / Q000235:genetics*; D004331:Drosophila melanogaster / Q000235:genetics*; D005786:Gene Expression Regulation; D018014:Gene Transfer Techniques; D017344:Genes, Insect; D012339:RNA, Ribosomal, 28S / Q000235:genetics*; D018626:Retroelements / Q000235:genetics*
D012339:RNA, Ribosomal, 28S; D018626:Retroelements