Three-Gene Combinations Involving Rph20 Provide Stable Resistance to Leaf Rust in Barley

Elite barley (Hordeum vulgare L.) breeding lines from the Northern Region Barley Breeding Program in Australia were evaluated at seedling and adult plant growth stages for leaf rust (LR; Puccinia hordei Otth) responses. The parental germplasm was obtained from different geographic origins and represented F3:5 lines spanning four years of trials (2009, 2011, 2012 and 2013). The 2009 and 2011 breeding populations (BPs) comprised a total of 360 lines and were genotyped with 3,244 polymorphic Diversity Arrays Technology (DArT) markers. The 2012 and 2013 BPs comprised 321 lines genotyped with the DArT genotyping-by-sequencing array (DArT-seq), which provided 15,400 polymorphic markers. Association mapping (AM) using the DArT/DArT-seq datasets and phenotypic data from 14 LR assays, identified 20 genomic regions associated with resistance. The Adult plant resistance (APR) locus Rph20 was the only region detected across all four BPs. Alignment of QTL across populations and previous studies identified three promising APR gene targets positioned on chromosomes 2H (at 22.17 cM), 4H (50.85 cM), and 7H (1.77 cM). Gene assignment using key marker haplotypes for the APR candidates and Rph20 revealed an additive effect for resistance to LR across environments and years. However, combining more than three genes failed to reduce mean disease response consistently. The frequency of APR gene combinations observed in breeding material suggests that three-gene combinations involving Rph20 were under selection. To achieve high levels of resistance to LR that are consistent across pathotypes and environments, we propose utilisation of Rph20 in combination with two additional APR genes.