Fine mapping of Rha2 in barley reveals candidate genes for resistance against cereal cyst nematode

Bart Van Gansbeke,John G Lewis,Kelvin H P Khoo,Diane E Mather,Kenneth J Chalmers

doi:10.1007/s00122-019-03279-3

Abstract

Key messageThe cereal cyst nematode resistance locus Rha2 was mapped to a 978 kbp region on the long arm of barley chromosome 2H. Three candidate genes are discussed.The cereal cyst nematode (CCN) Heterodera avenae is a soil-borne obligate parasite that can cause severe damage to cereals. This research involved fine mapping of Rha2, a CCN resistance locus on chromosome 2H of barley. Rha2 was previously mapped relative to restriction fragment length polymorphisms (RFLPs) in two mapping populations. Anchoring of flanking RFLP clone sequences to the barley genome assembly defined an interval of 5077 kbp. Genotyping-by-sequencing of resistant and susceptible materials led to the discovery of potentially useful single nucleotide polymorphisms (SNPs). Assays were designed for these SNPs and applied to mapping populations. This narrowed the region of interest to 3966 kbp. Further fine mapping was pursued by crossing and backcrossing the resistant cultivar Sloop SA to its susceptible ancestor Sloop. Evaluation of F2 progeny confirmed that the resistance segregates as a single dominant gene. Genotyping of 9003 BC2F2 progeny identified recombinants. Evaluation of recombinant BC2F3 progeny narrowed the region of interest to 978 kbp. Two of the SNPs within this region proved to be diagnostic of CCN resistance across a wide range of barley germplasm. Fluorescence-based and gel-based assays were developed for these SNPs for use in marker-assisted selection. Within the candidate region of the reference genome, there are nine high-confidence predicted genes. Three of these, one that encodes RAR1 (a cysteine- and histidine-rich domain-containing protein), one that is predicted to encode an acetylglutamate kinase and one that is predicted to encode a tonoplast intrinsic protein, are discussed as candidate genes for CCN resistance.

Highlights

The cereal cyst nematode (CCN) Heterodera avenae is a soil-borne parasite that infects the roots of many grass species and can cause significant yield losses in cereal crops including barley (Hordeum vulgare L.), wheat (Triticum aestivum L.) and oat (Avena sativa L.)
Phenotypic evaluation of a small panel of accessions in a tube test showed that Athinais, Bajo Aragon, Barley 191, Martin 403-2, Morocco, Morocco (Early), Nile, Orge Martin, Orge Martin 839, Sabarlis and Siri are all resistant to the H. avenae pathotype that was used here, with mean numbers of white cysts ranging from 0 to 1.1 per plant (Table S1 in Online Resource 1) while Alfa, Drost, Herta, Ortolan, Schooner and Varde are all susceptible to that pathotype, with mean numbers of white cysts ranging from 5.4 to 10.7 per plant
Sequences associated with five restriction fragment length polymorphisms (RFLPs) markers that Kretschmer et al (1997) had mapped near the resistance locus were anchored to the pseudomolecule sequence for chromosome 2H at positions between 654,782 and 684,123 kbp (Table S4 in Online Resource 1)

Summary

Introduction

The cereal cyst nematode (CCN) Heterodera avenae is a soil-borne parasite that infects the roots of many grass species and can cause significant yield losses in cereal crops including barley (Hordeum vulgare L.), wheat (Triticum aestivum L.) and oat (Avena sativa L.). Within host plant species that are affected by cyst nematodes, there is genetic variation for resistance (the ability to reduce nematode populations in the soil). In barley (reviewed below) and in wheat (reviewed by Jayatilake et al 2015), resistance loci have been genetically mapped and resistance alleles have been used in cereal breeding. In Australia, where there is thought to be only one pathotype (Ha13) of H. avenae, consistent use of resistant cereal cultivars and cultural management practices has been very effective in reducing nematode populations in agricultural soils and preventing yield losses (Murray and Brennan 2010)

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Conclusion