Identification and Differentiation of Clavibacter michiganensis Subspecies by Polymerase Chain Reaction-based Techniques

For the identi_cation and di}erentiation of the _ve sub! species of Clavibacter michi`anensis\ two di}erent poly! merase chain reaction "PCR#!based techniques were employed] ampli_cation with subspecies!speci_c primers and ampli_cation with random primers "RAPD#[ Based on the sequence data of the intergenic spacer region between 05S and 12S rRNA genes\ primers were designed for the identi_cation of each subspecies[ Using the designed primer pairs it was possible to identify each subspecies of C[ michi`anensis according to the ampli! _cation of a speci_c DNA fragment[ No ampli_cation products were obtained\ when bacteria belonging to other genera were submitted to PCR under the same conditions[ RAPD!PCR conditions suitable for the di}erentiation of C[ michi`anensis subspecies were developed[ RAPD typing was capable of distinguishing subspecies of C[ michi`anensis as well as strains within subspecies[ Both genomic variation between subspecies and genetic polymorphisms between bacterial strains were identi_ed as di}erences in the size and numbers of DNA fragments obtained