Figure 2.
Isolation of the bca gene replacement in GBS through homologous recombination. The shuttle/suicide vector pJL1147 was crossed into the wild-type strain A909. KmrCmr clones were selected at 37°C for sco mutants. The first recombination event occurs through homologous sequences in the flanking region of the bca gene resulting in JL2050. Interestingly, in all sco mutants mapped the crossover occurred in the N-terminal region. The second recombination event occurs between the C-terminal flanking duplicated regions created in sco mutants lead to excision of the integrated plasmid, thereby creating the dco mutant JL2053. The drawing is not to scale, but the HindIII and HaeIII sites used for mapping the mutants are shown; in addition, specific probes for the bca structural gene (A) and the N-terminal flanking region (B) used in Southern blots (Fig. 3) are illustrated.