Abstract
BackgroundRust fungi are an important group of plant pathogens that cause devastating losses in agricultural, silvicultural and natural ecosystems. Plants can be protected from rust disease by resistance genes encoding receptors that trigger a highly effective defence response upon recognition of specific pathogen avirulence proteins. Identifying avirulence genes is crucial for understanding how virulence evolves in the field.ResultsTo facilitate avirulence gene cloning in the flax rust fungus, Melampsora lini, we constructed a high-density genetic linkage map using single nucleotide polymorphisms detected in restriction site-associated DNA sequencing (RADseq) data. The map comprises 13,412 RADseq markers in 27 linkage groups that together span 5860 cM and contain 2756 recombination bins. The marker sequences were used to anchor 68.9 % of the M. lini genome assembly onto the genetic map. The map and anchored assembly were then used to: 1) show that M. lini has a high overall meiotic recombination rate, but recombination distribution is uneven and large coldspots exist; 2) show that substantial genome rearrangements have occurred in spontaneous loss-of-avirulence mutants; and 3) identify the AvrL2 and AvrM14 avirulence genes by map-based cloning. AvrM14 is a dual-specificity avirulence gene that encodes a predicted nudix hydrolase. AvrL2 is located in the region of the M. lini genome with the lowest recombination rate and encodes a small, highly-charged proline-rich protein.ConclusionsThe M. lini high-density linkage map has greatly advanced our understanding of virulence mechanisms in this pathogen by providing novel insights into genome variability and enabling identification of two new avirulence genes.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-3011-9) contains supplementary material, which is available to authorized users.
Highlights
Rust fungi are an important group of plant pathogens that cause devastating losses in agricultural, silvicultural and natural ecosystems
Genomic DNAs extracted from strains C and H, their F1 offspring (CH5) and 77 F2 progeny obtained by self-fertilisation of CH5 [48] were digested with PstI or NsiI and used to generate individually barcoded Restriction-site associated DNA sequencing (RADseq) libraries that were sequenced using 100 bp paired-end Illumina sequencing
The generation of a high-density genetic map has greatly improved the utility of the fragmented M. lini genome assembly and we have used these genetic resources together to identify two new avirulence genes by map-based cloning
Summary
Rust fungi are an important group of plant pathogens that cause devastating losses in agricultural, silvicultural and natural ecosystems. Plants can be protected from rust disease by resistance genes encoding receptors that trigger a highly effective defence response upon recognition of specific pathogen avirulence proteins. Rust fungi are an important group of plant pathogens that cause devastating diseases in agricultural, silvicultural and natural ecosystems. Five avirulence genes (AvrL567, AvrM, AvrP, AvrP123 and AvrP4) have been identified at four different loci in M. lini [16,17,18] They encode secreted proteins produced in a specialised fungal structure called the haustorium, which is a major site for signal exchange and nutrient acquisition in rust pathogens and other biotrophic fungi [19, 20]
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