Bioluminescence detection system of mutagen using firefly luciferase genes introduced in Escherichia coli lysogenic strain

1992 ◽  
Vol 64 (17) ◽  
pp. 1755-1759 ◽  
Author(s):  
Soo Mi. Lee ◽  
Masayasu. Suzuki ◽  
Michiyo. Kumagai ◽  
Hideo. Ikeda ◽  
Eiichi. Tamiya ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Detection System ◽  
Firefly Luciferase ◽  
Lysogenic Strain

Bioluminescence detection system of mutagen using firefly luciferase genes introduced in Escherichia coli lysogenic strain

1992 ◽  
Vol 64 (17) ◽  
pp. 833A-838A ◽  
Author(s):  
Soo Mi Lee ◽  
Masayasu Suzuki ◽  
Michiyo Kumagai ◽  
Hideo Ikeda ◽  
Eiichi Tamiya ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Detection System ◽  
Firefly Luciferase ◽  
Lysogenic Strain

scholarly journals Measurement of Effects of Antibiotics in Bioluminescent Staphylococcus aureus RN4220

2001 ◽  
Vol 45 (12) ◽  
pp. 3456-3461 ◽  
Author(s):  
Mervi Tenhami ◽  
Kaisa Hakkila ◽  
Matti Karp
Keyword(s):  
Staphylococcus Aureus ◽  
High Throughput Screening ◽  
Pathogenic Bacteria ◽  
Antimicrobial Agents ◽  
Light Emission ◽  
Detection System ◽  
Firefly Luciferase ◽  
Luciferase Reporter ◽  
Luciferase Expression ◽  
Luciferase Reporter Gene

ABSTRACT The spread of antibiotic resistance among pathogenic bacteria is a serious threat to humans and animals. Therefore, unnecessary use should be minimized, and new antimicrobial agents with novel mechanisms of action are needed. We have developed an efficient method for measuring the action of antibiotics which is applied to a gram-positive strain,Staphylococcus aureus RN4220. The method utilizes the firefly luciferase reporter gene coupled to the metal-induciblecadA promoter in a plasmid, pTOO24. Correctly timed induction by micromolar concentrations of antimonite rapidly triggers the luciferase gene transcription and translation. This sensitizes the detection system to the action of antibiotics, and especially for transcriptional and translational inhibitors. We show the results for 11 model antibiotics with the present approach and compare them to an analytical setup with a strain where luciferase expression is under the regulation of a constitutive promoter giving only a report of metabolic inhibition. The measurement of light emission from intact living cells is shown to correlate extremely well (r = 0.99) with the conventional overnight growth inhibition measurement. Four of the antibiotics were within a 20% concentration range and four were within a 60% concentration range of the drugs tested. This approach shortens the assay time needed, and it can be performed in 1 to 4 h, depending on the sensitivity needed. Furthermore, the assay can be automatized for high-throughput screening by the pharmaceutical industry.

scholarly journals Membrane-permeable luciferin esters for assay of firefly luciferase in live intact cells

1991 ◽  
Vol 276 (3) ◽  
pp. 637-641 ◽  
Author(s):  
F F Craig ◽  
A C Simmonds ◽  
D Watmore ◽  
F McCapra ◽  
M R H White
Keyword(s):  
Escherichia Coli ◽  
Mammalian Cells ◽  
Firefly Luciferase ◽  
Luciferase Expression ◽  
Intact Cells ◽  
Cos Cells ◽  
Escherichia Coli Cells ◽  
Luminescent Signal ◽  
The Difference ◽  
Kinetics Of

Five esters of luciferin were synthesized and compared with native luciferin as substrates for firefly luciferase expressed in live intact mammalian cells. The esters themselves were not substrates for purified luciferase, but four were substrates for a purified esterase and all appeared to be hydrolysed to luciferin within mammalian cells. At a substrate concentration of 0.01 mM, the peak luminescence from the cos cells expressing luciferase was up to 6-fold greater with the esters than with unmodified luciferin. At 0.1 mM, the difference between luciferin and the esters was decreased. The kinetics of the luminescent signal with the different luciferin esters varied significantly, indicating possible differences in the rates of uptake, breakdown and enzyme inhibition. The esters did not support luminescence from Escherichia coli cells expressing firefly luciferase, suggesting a lack of appropriate esterase activity in this particular strain. The esters could be useful for the assay of luciferase expression in intact mammalian cells when luciferin levels are limiting, for example in tissues, and in plants. Alternative luciferin derivatives may allow further improvements in sensitivity.

scholarly journals In Vivo Bioluminescence Imaging for the Study of Intestinal Colonization by Escherichia coli in Mice

2009 ◽  
Vol 76 (1) ◽  
pp. 264-274 ◽  
Author(s):  
M.-L. Foucault ◽  
L. Thomas ◽  
S. Goussard ◽  
B. R. Branchini ◽  
C. Grillot-Courvalin
Keyword(s):  
Escherichia Coli ◽  
Bioluminescence Imaging ◽  
Light Emission ◽  
Direct Method ◽  
Firefly Luciferase ◽  
Intestinal Colonization ◽  
Bacterial Populations ◽  
Bioluminescence Signal

ABSTRACT Bioluminescence imaging (BLI) is emerging as a powerful tool for real-time monitoring of infections in living animals. However, since luciferases are oxygenases, it has been suggested that the requirement for oxygen may limit the use of BLI in anaerobic environments, such as the lumen of the gut. Strains of Escherichia coli harboring the genes for either the bacterial luciferase from Photorhabdus luminescens or the PpyRE-TS and PpyGR-TS firefly luciferase mutants of Photinus pyralis (red and green thermostable P. pyralis luciferase mutants, respectively) have been engineered and used to monitor intestinal colonization in the streptomycin-treated mouse model. There was excellent correlation between the bioluminescence signal measured in the feces (R 2 = 0.98) or transcutaneously in the abdominal region of whole animals (R 2 = 0.99) and the CFU counts in the feces of bacteria harboring the luxABCDE operon. Stability in vivo of the bioluminescence signal was achieved by constructing plasmid pAT881(pGB2ΩPamiluxABCDE), which allowed long-term monitoring of intestinal colonization without the need for antibiotic selection for plasmid maintenance. Levels of intestinal colonization by various strains of E. coli could be compared directly by simple recording of the bioluminescence signal in living animals. The difference in spectra of light emission of the PpyRE-TS and PpyGR-TS firefly luciferase mutants and dual bioluminescence detection allowed direct in vitro and in vivo quantification of two bacterial populations by measurement of red and green emitted signals and thus monitoring of the two populations simultaneously. This system offers a simple and direct method to study in vitro and in vivo competition between mutants and the parental strain. BLI is a useful tool to study intestinal colonization.

scholarly journals Cloning of firefly luciferase cDNA and the expression of active luciferase in Escherichia coli.

1985 ◽  
Vol 82 (23) ◽  
pp. 7870-7873 ◽  
Author(s):  
J. R. de Wet ◽  
K. V. Wood ◽  
D. R. Helinski ◽  
M. DeLuca
Keyword(s):  
Escherichia Coli ◽  
Firefly Luciferase

scholarly journals Mechanism of phage production by a lysogenic strain of Escherichia coli

1953 ◽  
Vol 8 (2) ◽  
pp. 235-239
Author(s):  
Seijiro UCHIDA
Keyword(s):  
Escherichia Coli ◽  
Lysogenic Strain ◽  
Phage Production

scholarly journals Detection of Verotoxigenic Escherichia coli by Magnetic Capture-Hybridization PCR

1998 ◽  
Vol 64 (1) ◽  
pp. 147-152 ◽  
Author(s):  
Jinru Chen ◽  
Roger Johnson ◽  
Mansel Griffiths
Keyword(s):  
Escherichia Coli ◽  
Magnetic Beads ◽  
Detection System ◽  
Toxin Production ◽  
Enzyme Linked Immunosorbent Assay ◽  
Specific Pcr ◽  
Magnetic Capture ◽  
Pcr Products ◽  
Pure Cultures ◽  
Particle Separator

ABSTRACT Magnetic capture-hybridization PCR (MCH-PCR) was used for the detection of 36 verotoxigenic (verotoxin [VT]-producing)Escherichia coli (VTEC), 5 VTEC reference, and 13 non-VTEC control cultures. The detection system employs biotin-labeled probes to capture the DNA segments that contain specific regions of the genes for VT1 and VT2 by DNA-DNA hybridization. The hybrids formed were isolated by streptavidin-coated magnetic beads which were collected by a magnetic particle separator and, subsequently, amplified directly by conventional PCR. The detection system was found to be specific for VTEC: no amplification was obtained from non-VTEC controls, whereas VTEC isolates tested positive for one or two specific PCR products. With 5, 7, or 10 h of enrichment, the limits of detection were 103, 102, and 100 CFU/ml, respectively, by agarose gel electrophoresis. Southern hybridization did not seem to improve the limit of the detection. When applied to food, MCH-PCR was capable of detecting 100 CFU of VTEC per g of ground beef with 15 h of nonselective enrichment. The results of MCH-PCR for pure cultures of VT1- and/or VT2-producing E. coli cells were in total agreement with toxin production as measured by a VT enzyme-linked immunosorbent assay.

Enhanced Chloramphenicol Resistance of the Lysogenic Strain Escherichia Coli GY 2354 P1CM

1980 ◽  
pp. 153-155
Author(s):  
C. H. Hoffmeier ◽  
D. Noack ◽  
R. Geuther ◽  
H. Branä
Keyword(s):  
Escherichia Coli ◽  
Chloramphenicol Resistance ◽  
Lysogenic Strain

Photographic detection of luminescence in Escherichia coli containing the gene for firefly luciferase

1987 ◽  
Vol 161 (2) ◽  
pp. 501-507 ◽  
Author(s):  
Keith V. Wood ◽  
Marlene DeLuca
Keyword(s):  
Escherichia Coli ◽  
Firefly Luciferase
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