Apparent and real recombination frequencies in multicopy plasmids: the need for a novel approach in frequency determination.

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.

journal_name

J Bacteriol

journal_title

Journal of bacteriology

authors

Chédin F,Dervyn R,Ehrlich SD,Noirot P

doi

10.1128/jb.179.3.754-761.1997

subject

Has Abstract

pub_date

1997-02-01 00:00:00

pages

754-61

issue

3

eissn

0021-9193

issn

1098-5530

journal_volume

179

pub_type

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