Representational difference analysis to identify genomic differences between two related pathogenic strains of Pseudomonas aeruginosa

Representational difference analysis (RDA) is a recently developed technique used for amplifying genetic differences between two closely related genomes. The technique of RDA was adapted to the search for genes present in the PA14 strain but absent from the PA01 strain in Pseudomonas aeruginosa. Genes present in PA14 strain were physically subtracted from PA01 strain. The genes obtained from the second-round subtraction were tested by Southern blotting for their reactivity with the two species. The libraries achieved were comprehensive and specific in that they contain sequences corresponding to the presently identified PA14-specific genes. Of 16 randomly chosen clones specific to PA14 strain, the colony PCR confirmed that the majority of the products had the different size of genes. These results indicated that a number of unique fragments could be generated from the comparison of the two strains, that is consistent with the probable genetic differences between the two strains.