| Luciferase takes an important role in biochemical reaction of light-emitting in the microoganism, which involves the oxidation of reduced riboflacin phosphate and a long chain fatty aldehyde with the emission of the blue-green light. The combination of imaging technologies and luciferase-based bioluminescent bacterial reporter strains provide a sensitive and simple non-invasive detection method for the study of diverse biological processes, as well as in live animal models of disease.In this study, we transformed the pHSG396-luxAB plasmid carrying with lux operon into E.coli Top10 strain, and performed the biological characteristics of E. coli growth curve assay. It established the foundation for probiotic foods such as microbial in the application and provided a reference for study of enzymatic property and enzyme molecule modification.At last, the study prepared a pMOD–luxAB transposition, which take advantage of characteristics of stable highly expression for transposon, so it provided an original strain for research in environment and food industry.The results of the study were as follows:(1) Nucleic acid electrophoresis and SDS-PAGE indicated: recombinant plasmid was transformed successfully and the luciferase was expressed.(2) The optimum fermentation condition was: pH 6.0, the final concentration of capraldehyde 1.25 ‰, the amount of medium 50 m L/150 mL, inoculum size 1%, the final concentration of IPTG 0. 1 mmol/L, incubating at 37℃.(3) Results of orthogonal experimental design showed that factors that affected the size of light intensity were inoculum size, pH, temperature, the final concentration of IPTG and initial pH 6.0, the final concentration of IPTG 0.08 mmol/L, 1.2% and incubating at 36 ℃had the best expression of LUX, and the luminosity reached 1262.7.(4) At last, a pMOD –luxAB transposition complex was prepared successfully by means of cloning, ligation, translation and sequencing. |
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