Fig. 5.

Chromosomal organization of ppk1 ppx and construction/verification of the Δppk1::Tet mutation. In PAO1, ppk1 (PA5242; 2,210 bp) and ppx (PA5241; 1,520 bp) are convergently transcribed, and their ORFs overlap by 14 bp at the 3′ ends, denoted by the hatched area. Construction of the mutation is described in Materials and Methods. Tet denotes the tetracycline resistance-encoding gene cassette from pDMB100 (28). Ba and Bg indicate BamHI and BglII endonuclease restriction sites, respectively. The small arrows denote primers (described in SI Table 3) used in cloning ppk1 and in the genetic verification by PCR of the mutation (see Materials and Methods).