title: Amplification of a chimeric plasmid carrying an erythromycin-resistance determinant introduced into the genome of Streptococcus pneumoniae.
abstract: Hybrid plasmid molecules carrying an insert of pneumococcal DNA can integrate into the pneumococcal genome by homologous recombination. The resulting structure is a duplication of the pneumococcal DNA insert bracketing a vector genome. To select for plasmid integration, the vector plasmid was marked with an erythromycin (ery) resistance determinant (eryr) originating from pVA736, a streptococcal plasmid. Experiments with pR27 and pR28, two plasmids carrying the same insert of pneumococcal DNA but in opposite orientations, led to the following observation: (i) In one orientation of the ery region with respect to the amiA locus, cells exhibited a low-level resistance to ery; when these cells were grown in the presence of ery, amplification of the integrated plasmid occurred and cells became resistant to a high level of antibiotic. (ii) In the opposite orientation, a high level of resistance was observed, without need for amplification. These results indicate that, in the orientation conferring a high-level resistance without amplification, the ery region is transcribed both from its own promoter and from the promoter of the amiA locus. In the opposite orientation, a low level of transcription from the eryr promoter could account for a strong selective pressure for the amplified state, which then allows for rapid growth in the presence of ery.
mesh_terms: D004274:DNA, Recombinant; D004352:Drug Resistance, Microbial; D004917:Erythromycin / Q000494:pharmacology; D005784:Gene Amplification; D005798:Genes, Bacterial; D009876:Operon; D010957:Plasmids; D013296:Streptococcus pneumoniae / Q000187:drug effects / Q000235:genetics*
chemical_list: D004274:DNA, Recombinant; D004917:Erythromycin