title: [Study on gene knock-out in Mycobacterium BCG].
abstract: OBJECTIVE
  To establish the methodology of plasmid for gene knock-out in Mycobacterium BCG.
  
  
  METHODS
  We designed two pairs of primers for amplification of MDP1 gene and inserted two fragments into pKO plasmid, and then the recombinant plasmid for MDP1 gene knock-out was obtained, and named pKO-MDP1. Gene exchange took place within the genome of BCG after pKO-MDP1 plasmid was transformed into Mycobacterium BCG. The strain of Mycobacterium BCG with MDP1 gene knock-out was selected, and the curve of growth rate was studied.
  
  
  RESULTS
  The target strain was that of the positive strain by two step PCR and one step sucrose counter selection, without growth in culture media with hygomycin. The value of A(600) per 12 hours was detected for sixteen days. A "S" shaped growth curve was detected. However, there was no significant difference in the growth rates between the wild type Mycobacterium BCG strain and the gene knock-out Mycobacterium BCG strain.
  
  
  CONCLUSIONS
  The plasmid of pKO was a useful tool for gene knock-out in Mycobacteria. MDP1 maybe one of the factors influencing the growth rate, but it was not the only one.
authors.affiliation: Department of Bacteriological Immunology, Beijing Tuberculosis & Thoracic Tumour Research Institute, Beijing 101149, China.
authors.forename: Xiao-you
authors.initials: XY
authors.lastname: Chen
authors.forename: Chuan-you
authors.initials: CY
authors.lastname: Li
authors.forename: Yu
authors.initials: Y
authors.lastname: Ma
authors.forename: Chong
authors.initials: C
authors.lastname: Liu
authors.forename: Jing-hui
authors.initials: JH
authors.lastname: Wang
authors.forename: Xue-feng
authors.initials: XF
authors.lastname: Zhang
authors.forename: Zeng-yi
authors.initials: ZY
authors.lastname: Chang
mesh_terms: D001500:BCG Vaccine; D001426:Bacterial Proteins / Q000235:genetics* / Q000302:isolation & purification; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000302:isolation & purification; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D009163:Mycobacterium bovis / Q000235:genetics*; D009169:Mycobacterium tuberculosis / Q000235:genetics*; D010957:Plasmids; D011995:Recombination, Genetic; D014169:Transformation, Bacterial
chemical_list: D001500:BCG Vaccine; D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C403010:MDP1 protein, Mycobacterium