PCR Assays for Detection of Pseudomonas tolaasii and Pseudomonas agarici

ABSTRACT

PCR assays were developed to detect Pseudomonas tolaasii and Pseudomonas agarici using primer sets, PTOF/PTOR and PAGF/R23-1R. The primer set, PTOF and PTOR, was designed from the nucleotide sequence of pPTOF2 that showed specificity for P. tolaasii in dot blot previously. For development of primers specific for P. agarici, ITS I regions of seven P. agarici strains were analyzed. P. agarici strains contained from one up to three putative ITS I regions, which were different in size and nucleotide sequence from each other. From the sequence of the band (PaI-III) common to all P. agarici strains, primer PAGF was designed. PAGF was used for forward primer, and R23-1R as reverse primer to detect P. agarici. Multiplex PCR with two primer sets, PTOF/PTOR and PAGF/R23-lR, successfully produced two fragments (PTSF and PASF) specific for P. tolaasii and P. agarici with the mixture of DNA of P. tolaasii and P. agarici.