Differentiation and Detection of the Subspecies of Clavibacter Michiganensis by PCR (Polymerase Chain Reaction) - Techniques
The five phytopathogenic subspecies of Clavibacter michiganensis (C. m. sepedonicus, C. m. michiganensis, C. m. insidiosus, C. m. tessellarius, C. m. nebraskensis) are causal agents of various diseases in agriculture. For the detection of each of these five subspecies a specific polymerase chain reaction (PCR)-assay was established. PCR primers were selected targeting genomic DNA. To obtain these primers, the spacer region between the 16S and 23S rRNA genes of these five subspecies was amplified and sequenced. On the basis of these sequence data primer pairs for each of the five subspecies were designed. The primer pairs (PSA 1–5) amplified a specific DNA fragment respectively only at that subspecies they were designed for and not at other related subspecies. The size of the amplified specific DNA fragment was for C. m. sepedonicus 502 bp (PSA-1), for C. m. michiganensis 270 bp (PSA-2), for C. m. insidiosus 393 bp (PSA-3), for C. m. tessellarius 587 bp (PSA-4) and for C. m. nebraskensis 393 bp (PSA-S).