Barley allele-specific amplicons useful for identifying wheat-barley recombinant chromosomes

Barley (Hordeum vulgare L.) is potentially a new source of genes for wheat (Triticum aestivum L.) improvement. Wheat-barley chromosome recombinant lines provide a means for introgressing barley genes to wheat genome by chromosome engineering, and since these are expected to occur only rarely in special cytogenetic stocks, an efficient selection skill is necessary to identify them. To convert RFLP markers to barley allele-specific PCR markers useful for effective production of wheat-barley recombinant lines, 91 primer sets derived from RFLP clones which were previously mapped to the barley chromosomes were examined for PCR amplification using 'Chinese Spring' wheat, 'Betzes' barley and the wheat-barley chromosome addition lines. The polymorphisms were detected by an agarose gel electrophoresis of the PCR products without digestion with restriction enzymes. Out of 81 primer sets producing polymorphisms between the wheat and barley genomes, 26 amplified barley chromosome-specific DNAs which were confirmed to be located on the same chromosome as the RFLP markers by using the wheat-barley chromosome addition lines. These amplified DNAs represent barley allele-specific amplicons, which distinguish barley alleles from their wheat homoeologous counterparts. The present investigation revealed a higher probability for obtaining allele-specific amplicons from genomic DNA-derived RFLP markers than from cDNA-derived ones. The barley allele-specific amplicons developed in this study, namely, four for chromosome 2H, two for 3H, seven for 4H, eight for 5H, one for 6H and four for 7H, are suitable for identifying 'Chinese Spring' wheat- 'Betzes' barley recombinant chromosomes. However, one out of eight barley allele-specific amplicons on chromosome 5H did not detect a unique barley band in a 'New Golden' barley chromosome 5H addition line of 'Shinchunaga' wheat, indicating there may be a need to reconstruct allele-specific amplicons with different barley cultivars.