Use of ccSSR markers for the determination of the purity of alloplasmic wheat in different Hordeum cytoplasms

We tested three different consensus chloroplast simple sequence repeat (ccSSR) primers to identify amplified polymorphic products in Hordeum chilense, Hordeum vulgare and other Triticeae species with a double aim. First, to use chloroplast-specific primers as an indirect method for the assessment of wheat cytoplasmic male sterile purity in seedlings during multiplication and in the development of alloplasmic lines. Second, to analyse cytoplasmic relationships among H. chilense accessions and between H. chilense and other members of the Triticeae tribe. The products from primer ccSSR-4 were easily discriminated using agarose gel electrophoresis. Based on the lengths of amplification products, three groups were identified: the first included almost exclusively H. chilense accessions, the second contained H. vulgare accessions and the third comprised the wheat and the rest of the analysed accessions. Sequencing of PCR products revealed point mutations and insertions/deletions in addition to the expansion/contraction of the microsatellite repeat length. Data analyses of sequenced fragments revealed six groups of accessions among the material studied. No significant differences were found among H. chilense accessions.