Development of molecular RAPD marker for the identification of Pediococcusacidilactici strains

A RAPD analysis performed using a single primer targeted to the pediocin Ac H/PA-1 gene was carried out on several P. acidilactici strains and on some related species of lactic acid bacteria. The high degree of genetic variabi lity detected in P. acidilactici strains did nor allow the selection of a c ommon RAPD fragment that could be chosen as a potential species-specific DN A marker. Nevertheless a 700 bp fragment, that was found to be peculiar of all potential pediocin producer strains analyzed, was cloned and sequenced with the aim to develop a species specific PCR marker. Sequence analysis of the cloned 700 bp fragment showed one putative small open reading frame (O RF1), with no significant homology with known genes, and a partial putative second coding region (ORF2) with a high degree of similarity with several methionyl tRNA synthetasis (metS) genes. The two coding regions were separa ted by a short spacer region. Primers targeted to ORF2 plus part of the spa cer region and primers designed for the amplification of the entire cloned RAPD fragment were found to be species-specific for the detection of P. aci dilactici strains. Furthermore primers designed on the ORF1 sequence allowe d the amplification of a 439 bp fragment only in some P. acidilactici strai ns, including pediocin producing strains.