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Construction of stable, single-copy luciferase gene fusions in Escherichia coli

Archives of Microbiology, ISSN: 0302-8933, Vol: 156, Issue: 6, Page: 444-448
1991
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A ColE1-based plasmid for transcriptional gene fusions was constructed that contains both the promoterless luxAB genes of Vibrio harveyi and a tet marker within the inverted repeats of a left end-truncated Tn 5 element. Introduction of this plasmid into an Escherichia coli strain containing a plasmid (pTF421) that overproduces ColE1 RNA1 (and thus inhibits replication of the ColE1 plasmid) allowed selection for cells that had a single copy of the luxAB operon transposed into the chromosome beginning 5 days post-transformation. The long latent period necessary for Tn 5 transposition is analogous to that found in other systems, where transposition frequencies and mutation rates increase in a time-dependent manner when selected for upon prolonged incubation on Petri dishes under bacteriostatic conditions. © 1991 Springer-Verlag.

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