Sensitive biological detection method for tetracyclines using a tetA-lacZ fusion system.
A sensitive microbiological detection system for tetracyclines, utilizing an Escherichia coli strain containing a cloned tetA-lacZ gene fusion, is described. Expression of beta-galactosidase by the fusion plasmid pUB3610 remained subject to regulatory control by the TetR repressor protein, with the presence of tetracyclines in the growth medium leading to a 12-fold induction of beta-galactosidase synthesis. Because synthesis of beta-galactosidase was influenced to a small extent by the carbon source and the addition of cyclic AMP to the medium, cells were grown in the presence of cyclic AMP to enhance the sensitivity of the assay. All commonly marketed tetracyclines and some derivatives at concentrations as low as 0.1 ng/ml could be detected in the growth medium. A plate assay utilizing the fusion plasmid that detects 1 ng of tetracycline has also been developed.
Chopra
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I
Department of Microbiology, School of Medical Sciences, University of Bristol, United Kingdom.
Hacker
K
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Misulovin
Z
Z
Rothstein
D M
DM
D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D003470:Culture Media; D000242:Cyclic AMP / Q000494:pharmacology; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D005696:Galactosidases / Q000096:biosynthesis; D005798:Genes, Bacterial; D013753:Tetracycline Resistance; D013754:Tetracyclines / Q000032:analysis* / Q000494:pharmacology
D003470:Culture Media; D013754:Tetracyclines; D000242:Cyclic AMP; D005696:Galactosidases