Figure 6: AgeI restriction analysis of clones derived from cointegrants during homologous recombination. (a) Cointegrants formed from the first stage of RecA-mediated recombination underwent a second recombination event to remove the integrated pKOV Kan plasmid. This second recombination event could result in either a revertant or a mutant genotype. Of the 12 colonies screened, clone 9D had the desired restriction pattern consistent with a mutated ORF51 gene. (b) The mutant clone 9D was digested and run alongside a revertant 3A and the cointegrant clone 9 from which it originated. The loss of the integrated pKOV Kan plasmid can be visualised by the 52 kb band in the final lane decreasing in size by 7 kb to 45 kb.