Screening and confirmation of pta and ack genes knockout. (A) polymerase chain reaction (PCR) screening using genomic DNA as template. Lane 1, 1 kb DNA ladder (TaKaRa), Lane 2, SCUT27, Lane 3&4, positive isolates. (B) Southern blot analysis of genomic DNA from wild type SCUT27 (Lane 1) and the pta and ack deletion clones, LA1002 (Lane 2) digested with Pst I. The probe with the expected sizes of 486 bp (position shown in Figure 1), hybridized to one 1.2 kb fragment of wild type DNA, and to one 2.2 kb fragment of the mutant DNA.