Limited use of the Cre/loxP recombination system in efficient production of loxP-containing minicircles in vivo.
The Cre/loxP recombination system of bacteriophage P1 is one of the most powerful tools in genome engineering. We report, however, that the activity of the Cre/loxP system interferes with the stability of the multicopy loxP-bearing plasmids in Escherichia coli recA bacteria. Due to the predominantly unidirectional Cre-mediated high-order multimer formation of these plasmids, the number of their copies (overall yield) gradually decreases. Intermolecular recombination reduces the copy number of plasmids and eventually increases their segregational instability. We have found that in the presence of even the slightest amount of Cre activity, loxP-bearing plasmids continuously undergo multimerization, which very rapidly leads to loxP-plasmid free cells. Our results are compatible with the hypothesis of the multimer catastrophe [Cell, 1984 (36), 1097].
Sektas
Marian
M
Department of Microbiology, University of Gdansk, 80-822 Gdansk, ul.Kladki 24, Poland. sektas@biotech.univ.gda.pl
Specht
Maciej
M
D017101:Bacteriophage P1 / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development; D005818:Genetic Engineering / Q000379:methods*; D019426:Integrases / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011938:Rec A Recombinases / Q000235:genetics / Q000378:metabolism; D011995:Recombination, Genetic; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism
D014764:Viral Proteins; C045073:Cre recombinase; D019426:Integrases; D011938:Rec A Recombinases