Fig. 3

PCR validation of DFCT13, an rsbv1 insertion mutant created using the GII(bla) ampicillin-selection cassette. EBs were transformed with pDFTT6bla and serially passaged with ampicillin selection. Mutants were plaque purified and a single plaque was expanded for PCR and phenotype analysis (Figs. 5, 6). The insertion locus map is shown in panel a with the intron highlighted in pink. The intron is inserted in an anti-sense orientation to rsbV1. DNA was isolated from EBs and used for PCR. Expected product sizes are listed in panel b and the primers used along with reaction notes are listed in Table 1. PCR products were separated and visualized as described for Fig. 2. PCR results are shown for the wild type strain, DFCT13, and pDFTT6bla in panels c, d, and e, respectively