Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving viable micro-organism
Patent
1996-05-13
1998-08-04
Elliott, George C.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving viable micro-organism
435 4, 4352523, C12Q 102, C12N 121, C12N 100
Patent
active
057891885
ABSTRACT:
A method for detecting a tetracycline efflux pump inhibitor in the presence of tetracycline using a reporter gene system where the tetA promoter directs transcription of a reporter gene (lacZ) while the tetA is under the control of the tet repressor encoded by the tetR gene is described. The method uses a cell having a reporter gene system where the tetA promoter directs transcription of a reporter gene (lacZ) and an active efflux system in which relatively modest levels of the efflux protein encoded by the tetA gene are produced in a constitutive manner, i.e., not under the control of the tet repressor encoded by the tetR gene. Test samples which are inhibitors of the TetA efflux protein will allow accumulation of tetracycline inside the cells at levels which will induce expression of the tetA-lacZ transcriptional fusion to give a positive signal. A microorganism is also described which is refractory to induction by DNA damaging agents and comprises (1) an indicator gene fused to a tetA promoter, as a single- or low-copy number gene; (2) a tetR gene expressed at low levels, preferably at a level producing a sensitivity to 10 ng or less of tetracycline, and (3) a constitutive gene encoding a tetracycline efflux pump.
REFERENCES:
patent: 4806529 (1989-02-01), Levy
patent: 4911924 (1990-03-01), Griffith et al.
patent: 5001230 (1991-03-01), Brown et al.
McMurray, L.M. et al., "Active efflux of tetracycline encoded by four genetically different tetracycline resistance elements in Escherichia coli", Proc. Natl. Acad. Sci., 77:3974-3977, 1980.
Mojumdar, M. and S.A. Khar, Characterization of the tetracycline resistance gene of plasmid pT181 of Staphylococcus aureus. J. Bacteriol. 170:5522-5528, 1988.
Coleman, D.C. and T.J. Foster, "Analysis of the reduction in expression of tetracycline resistance determined by transposon Tn10 in the multicopy state", Mol. Gen. Genet. 182:171-177, 1981.
Moyed, J.S., et al., "Multicopy Tn10 tet plasmids confer sensitivity to induction of tat gene expression", J. Bacteriol., 155:549-556, 1983.
Bertrand, K.P., et al., "Overlapping divergent promoters control expression of Tn10 tetracycline resistance", Gene, 23:149-156, 1983.
Hillen, W. et al., "Control of expression of the Tn10-encoded tetracycline resistance operon II. Interaction of RNA polymerase and TET repressor with the tet operon regulatory region", J. Mol. Biol., 172:185-201, 1983.
Eckert, B. et al., "Overproduction of Transposon Tn10-Encoded Tetracycline Protein Results in Cell Death and Loss of Membrane Potential", J. Bact. 171:3557-3559, 1989.
Bertrand, K.P. et al., "Construction of a single-cop promoter vector and its use in analysis of regulation of the transposon Tn10 tetracycline resistance determinant", J. Bacteriol., 158:910-919, 1984.
Miller, J.H., "Experiments in Molecular Genatics", Cold Spring Harbor Laboratories, Cold Spring Harbor, N.Y.
Chopra, I. et al., "Sensitive biological detection method for tetracycline using a tetA-lacZ fusion system", Antimicrob. Agents Chemoth., 34:111-116, 1990.
Rothstein, D.M. et al., "Regulation of expression from the glnA promoter of Escherichia coli in the absence of glutamine synthetase", Proc. Nat. Acad. Sci. 77:7372-7376, 1980.
Hershey, A.D. (ed.), "The Bacteriophage Lamdda", Cold Spring Harbor Laboratories, Cold Spring Harbor, N.Y., 1971.
Kunkel, T.A. et al., "Rapid and efficient site-specific mutagenesis without phenotypic selection", Methods Enzymology, 154:367, 1987.
Davis, R.W. et al., "Advanced Bacterial Genetics", Cols Spring Harbor Laboratory, Cold Spring Harbor, N.Y., 1980.
Osburne, M.O., "In vivo inhibition of bacterial DNA gyrase by cinodinne, a glycocinnamoylspermidine antibiotic", Antimicrobial Agents and Chemotherapy, 34:1450-1452, 1990.
Smith, L.D. et al., "Mutations in the Tn10 repressor that interfere with induction, location of the tetracycline-binding domain", J. Mol. Biol., 203:929-959, 1988.
Bochner, B.R. et al., "Positive selection for loss of tetracycline resistance", J. Bacteriol, 143:926-933, 1980.
Schwyn, R. et al., "Universal chemical assay for the detection and determination of siderophores", Analytical Biochemistry, 160:47-56, 1987.
Gold, L. et al., "Escherichia coli and Salmonella Typhimutium: Cellular and Molecular Biology", F.C. Niedhard (ed.), American Society of Microbiology, Washington, D.C., vol. 2:1302-1307, 1987.
Dosch, D.C. et al., "Tetracycline Resistance Element of pBR322 Mediates Potassium Transport", J. Bacteriol. 160:1188-1190, 1984.
Rubin, R.A. and S. Levy, "Interdomain Hybrid Tet Proteins Confer Tetracycline Resistance Only When They are Derived from Closely Related Members of the tet Gene Family", J. Bacteriol., 172:2303-2312, 1990.
Bailey, J. E. (1991) Toward A Science of Metabolic Engineering. Science, vol. 252, pp. 1668-1675.
Guay Gordon Gerald
Rothstein David Michael
American Cyanamid Company
Barnhard Elizabeth M.
Elliott George C.
Lowney Karen A.
Railey II Johnny F.
LandOfFree
Tetracycline-efflux pump inhibitor screening methods does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Tetracycline-efflux pump inhibitor screening methods, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Tetracycline-efflux pump inhibitor screening methods will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-1175778