Confirmation of the presence of pYV in the original strains, cells in red pinpoint colonies, and cells in the white border around a red pinpoint colony from CR-MOX by PCR assay targeting a key regulatory gene virF, which encodes a transcriptional activator for the expression of pYV-encoded outer membrane protein Yop51. The primer pairs (5′-TCATGGCAGAACAGCAGTCAG-3′ and 5′-ACTCATCTTACCATTAAGAAG-3′) for detection of the virF gene (430- to 1020-nucleotide region) amplified a 591 base pair (bp) product from the virulence plasmid. Lane M, 50–1,000 bp ladder marker; lanes, 1, 5, and 9 showing the absence of 591-bp product in cells of the white borders of Y. enterocolitica, Y. pseudotuberculosis, and Y. pestis, respectively; lanes 2, 6, and 10 showing the presence of 591-bp product in the original strains of Y. enterocolitica, Y. pseudotuberculosis, and Y. pestis respectively before phenotypic evaluation; lanes 3, 7, and 11 showing the presence of 591-bp product in cells of the red pinpoint colonies of Y. enterocolitica, Y. pseudotuberculosis, and Y. pestis, respectively, and lanes 4, 8, and 12 showing the presence of 591-bp product within cells of red pinpoint colonies surrounded by white border of Y. enterocolitica, Y. pseudotuberculosis, and Y. pestis respectively [7].