Pulse-Field gel-electrophoretic analysis of the amplification and copy-number stability of an integrational plasmid in Bacillus subtilis |
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Authors: | C Vázquez-Cruz J C Ochoa-Sánchez G Olmedo-Alvarez |
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Institution: | (1) Departamento de Ingeniería Genética, CINVESTAV IPN, Unidad Irapuato Irapuato, Gto., México,;(2) Apartado Postal 629 Irapuato, Gto. 36500, México. Fax: (462) 4458-49 & 458-46 E-mail: golmedo@irapuato.ira.cinvestav.mx, |
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Abstract: | The stability of integration and amplification of an integrational plasmid in Bacillus subtilis was analyzed. A cat-containing plasmid was constructed that could be integrated into the amy locus to facilitate measurement of excision events. Pulse-field gel electrophoresis was used to measure the copy number in
strains that were resistant to different levels of chloramphenicol. The stability of the amplified unit in strains containing
from 2 to 18 tandem copies of the amplicon in the presence and absence of chloramphenicol and through different generation
times was then determined. Our results demonstrate that, for any given strain, the copy number of the amplicon remains stable.
Furthermore, this stability is maintained when a clone containing an amplicon of defined size is cultured through as many
as 100 generations in the absence of selective pressure.
Received: 27 October 1995/Received revision: 3 February 1996/Accepted: 11 March 1996 |
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