Apparent and real recombination frequencies in multicopy plasmids: the need for a novel approach in frequency determination.
J Bacteriol
; 179(3): 754-61, 1997 Feb.
Article
en En
| MEDLINE
| ID: mdl-9006030
ABSTRACT
Recombination studies of bacteria are often carried out with multicopy plasmids, and recombination frequencies are often deduced from the proportion of cells in the population that express a recombinant phenotype. These frequencies should however be called apparent frequencies, since detection of the recombinant cells requires not only the formation of a rearranged plasmid but also its establishment in the cell. The establishment of the recombinant plasmid can possibly be affected by its interaction with the parental plasmids. To test this hypothesis, we have used a plasmid system enabling the study of deletion formation between short direct repeats (18 bp) in Bacillus subtilis and developed a method by which deletion frequencies are measured under conditions under which interaction is abolished. Real deletion frequencies were thus determined and compared with apparent deletion frequencies. Real frequencies were underestimated by a factor ranging from 4- to 500-fold, depending upon the plasmid under study. This implies that a large majority of the recombinant molecules that are formed are generally not detected. We show that apparent deletion frequencies strongly depend upon (i) the parental plasmid copy number, (ii) the ability of the recombinant molecules to form heterodimeric plasmids, and (iii) the fitness of the recombinant molecules relative to that of parental molecules. Finally, we show that under conditions under which all recombinant molecules are scored, transcription can inhibit the deletion process 10-fold.
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Plásmidos
/
Recombinación Genética
/
Bacillus subtilis
/
Técnicas Genéticas
Idioma:
En
Revista:
J Bacteriol
Año:
1997
Tipo del documento:
Article
País de afiliación:
Francia