title: [Construction of luxAB-labelled Pseudomonas aeruginosa].
abstract: OBJECTIVE
  To construct a genetically engineered Pseudomonas aeruginosa expressing luxAB luciferase gene.
  
  
  METHODS
  Use the Bgl II enzyme to cut the plasmid pUC-luxAB, then recovered luxAB segment, and concatenate the plasmid carrier cut by the enzyme BamH I to form the reconstructed plasmid pBBR-luxAB, then transform E. coli DH5alpha competent cells. The positive clones that contain the reconstructed plasmid pBBR-luxAB experience the multiple screening by gentamycin (Gm), chloromycin (Cm) and luminescence detection, and at the same time, set up the control bacteria. Extract the plasmid pBBR-luxAB, then cut by enzyme, and then gel electrophoresis, and validate the rightness of the plasmid construction. By the mean of 2-generation mating, introduce the plasmid pBBR-luxAB into Pseudomonas aeruginosa, construct the gene engineered bacteria P. aeruginosa (pBBR-luxAB), and then measure the reproductive stableness of the plasmid, bioluminescence kinetic curves and relationship between luminescence and plate counting.
  
  
  RESULTS
  Successfully construct the pBBR-luxAB reconstructed plasmid, and surely, it is successfully introduced to P. aeruginosa. After 4 successively transfer, the pBBR-luxAB retaining rate still can keep at 93%. After the addition of substrates for 20 min, the luminous intensity of the reconstructed bacteria tends to be stable (1.32 mV/ml). There is a significantly positive correlation between luminous intensity and viable cell counting (r = 0.96, P < 0.05).
  
  
  CONCLUSIONS
  This research successfully constructed luxAB labelled P. aeruginosa, providing a useful means to study the survival, migration and damage of P. aeruginosa.
authors: Tianjin Centers for Disease Control and Prevention, Tianjin 300011, China. liwen_630@163.com
authors: Wen
authors: W
authors: Li
authors: Hongyan
authors: H
authors: Liu
authors: Zheng
authors: Z
authors: Huang
mesh_terms: D004269:DNA, Bacterial / Q000235:genetics; D004274:DNA, Recombinant / Q000235:genetics; D017930:Genes, Reporter / Q000235:genetics; D005818:Genetic Engineering / Q000379:methods*; D005819:Genetic Markers; D005822:Genetic Vectors; D008156:Luciferases / Q000235:genetics*; D008163:Luminescent Measurements; D011550:Pseudomonas aeruginosa / Q000235:genetics* / Q000502:physiology
chemical_list: D004269:DNA, Bacterial; D004274:DNA, Recombinant; D005819:Genetic Markers; D008156:Luciferases