FIG. 2.
Genomic locus of M. tuberculosis encompassing narGHJI and Southern blot analysis of narG mutants of M. tuberculosis, M. bovis, and M. bovis BCG. An EcoRV fragment of M. tuberculosis contains the entire narGHJI gene cluster, including 0.9-kb prior to the translation start of narG. MscI/BsmI restriction sites for the construction of the narG deletion in M. tuberculosis, and ClaI restriction sites for construction of narG deletions in M. bovis and M. bovis BCG are depicted. The positions of the deleted fragments are shown (ΔM. tuberculosis and ΔM. bovis, BCG). The positions of the forward primer and the reverse primer that were used for screening narG mutants are depicted as arrows. A Southern blot analysis showing wild-type (lanes 1 to 3) and mutant (lanes 4 to 6) strains is shown below. Genomic DNAs from M. tuberculosis (lane 1), M. bovis (lane 2), M. bovis BCG (lane 3), ΔnarG M. tuberculosis (lane 4), ΔnarG M. bovis (lane 5), and ΔnarG M. bovis BCG (lane 6) were digested with SmaI. The position of the DNA probe for Southern blot analysis is shown. The loss of a SmaI site due to deletion within the narG gene results in an “upshift” of the specific band in all three mutant strains.