Construction of mini-Tn10luxABcam/Ptac-ATS and its use for developing a bacteriophage that transduces bioluminescence to Escherichia coli O157:H7.
Mini-Tn10luxABcam/Ptac-ATS was constructed in order to develop a luciferase-transducing bacteriophage for detecting Escherichia coli O157:H7. The transposon was designed to deliver a 3.6-kb insertion that confers n-decanal-dependent bioluminescence and resistance to chloramphenicol and was constructed using mini-Tn10cam/Ptac-ATS in the plasmid pNK2884 and luxAB from Vibrio harveyi. PhiV10, a temperate bacteriophage infecting common phage types of Escherichia coli O157:H7, was mutagenized as a prophage in E. coli O157:H7 strain R508. PhiV10::luxABcamA1-23 was rescued from the strain by propagating it on a strain lacking the bacteriophage and the vector containing the transposon. The bacteriophage transduced n-decanal-dependent bioluminescence to E. coli O157:H7 strain R508 that was measurable approximately 1 h post infection.
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Health Canada, Health Protection Branch, Guelph Laboratory, 110 Stone Road West, Guelph, Ont., Canada. tom_waddell@hc-sc.ca
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D003090:Coliphages / Q000235:genetics* / Q000502:physiology; D004251:DNA Transposable Elements; D019453:Escherichia coli O157 / Q000235:genetics* / Q000502:physiology / Q000821:virology; D008156:Luciferases / Q000235:genetics; D008163:Luminescent Measurements; D008854:Microscopy, Electron; D016254:Mutagenesis, Insertional; D014161:Transduction, Genetic
D004251:DNA Transposable Elements; D008156:Luciferases