Simple sequence repeats in Puccinia graminis: abundance, cross-formae specialis and intra-species utility, and development of novel markers

We evaluated the abundance and nature of simple sequence repeats (SSRs) in the causal agent of stem rust of wheat, Puccinia graminis f. sp. tritici, in order to characterize and develop SSR markers. A strategy was tested in which publicly available genome datasets extracted from genome sequences of P. graminis f. sp. tritici were searched for abundance and mined for candidate novel SSR markers that could be applied to a wide range of Puccinia species. The genome analysis from 393 supercontigs yielded a total of 34,359 SSR repeat motifs. A majority of these (98 %) were mononucleotide, dinucleotide, and trinucleotide repeats. The AG/GA dinucleotide motifs were the most common, and a TTGTT motif was the longest of all SSRs. From a total of 500 primer pairs designed, 460 produced bands within the expected size range and 56 SSR markers were eventually developed. Most of these were tetranucleotide and pentanucleotide motifs with PIC values ranging from 0.40 to 0.91 with an average PIC value of 0.71. The cross amplification of these markers in 11 isolates representing five Puccinia species and six formae speciales of P. graminis was also investigated. The frequency of PCR amplifications was lower for the species P. coronata, P. striiformis and P. triticina, and consequently, no significant polymorphisms were detected. In contrast 10, 10, and 11 polymorphic SSR loci were identified in P. graminis f. sp. secalis, P. graminis f. sp. avenae, and the “Scabrum” rust, respectively. The SSR markers reported here will be useful to characterize and profile global diversity in the important pathogen P. graminis