High recombination rate of an Epstein-Barr virus-simian virus 40 hybrid shuttle vector in human cells.
The stability of an Epstein-Barr virus (EBV)-simian virus 40 (SV40) hybrid shuttle vector, the p205-GTI plasmid, was analyzed in human cells during EBV- or SV40-type replication mode. When the p205-GTI plasmid was maintained as an episomal EBV vector in the human 293 cell line, no rearrangement was detected. To induce the SV40 replication mode, cells containing the episomal p205-GTI plasmid were either transfected with vectors carrying the T antigen gene or infected with SV40. Surprisingly, we observed both production and amplification of different classes of recombinant molecules. Particular types of modifications were found in most of the recombinants. The most striking rearrangement was a duplication of the promoter and enhancer regions of SV40 which was inserted in the thymidine kinase (TK) promoter. This recombination process involved a few bases of homology, and one of the recombination junctions implicated the GC boxes which constitute the essential components of the TK and SV40 early promoters. Our results suggest that a combination of a low level of base homology and a specific DNA sequence function (promoter and enhancer sites) leads to a very high level of recombinational activity during T-antigen-dependent plasmid replication.
Stary
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Laboratory of Molecular Genetics, Institut de Recherches Scientifiques sur le Cancer, Villejuif, France.
James
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Sarasin
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D001483:Base Sequence; D002478:Cells, Cultured; D005784:Gene Amplification; D005822:Genetic Vectors; D004854:Herpesvirus 4, Human / Q000235:genetics*; D006801:Humans; D010957:Plasmids; D011995:Recombination, Genetic; D013539:Simian virus 40 / Q000235:genetics*; D014158:Transcription, Genetic; D014162:Transfection