PhiC31-mediated cassette exchange into a bacterial artificial chromosome

The use of bacterial artificial chromosomes (BACs) modified via homologous recombination in Escherichia coli has become a powerful tool in the transgenic field. Homologous recombination allows the manipulation of BACs in very different ways. However this process can be cumbersome and problematic whe...

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Bibliographic Details
Published inBioTechniques Vol. 43; no. 5; p. 659
Main Authors Blaas, Leander, Musteanu, Monica, Zenz, Rainer, Eferl, Robert, Casanova, Emilio
Format Journal Article
LanguageEnglish
Published England 01.11.2007
Subjects
Bacteriophages - enzymology
Chromosomes, Artificial, Bacterial - genetics
Integrases - metabolism
Mutagenesis, Insertional - methods
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Summary:The use of bacterial artificial chromosomes (BACs) modified via homologous recombination in Escherichia coli has become a powerful tool in the transgenic field. Homologous recombination allows the manipulation of BACs in very different ways. However this process can be cumbersome and problematic when using large targeting constructs containing several repeated elements. In order to address this problem, we have established a phiC31 integrase-mediated cassette exchange into a BAC. As an example of this technique, we have exchanged a cassette previously recombined into a BAC containing the Rosa 26 locus, by a 16.5-kb incoming construct containing several repeated elements. The combination of homologous recombination in E. coli and cassette exchange should expand the tools for manipulating BACs, thus facilitating the generation of constructs with higher complexity.
ISSN:0736-6205
1940-9818