Nat. Biotechnol. 37, 139–143 (2019).

Wild germplasm of crops contains abundant, yet underexplored, disease resistance (R) gene resources. The application of existing methods for cloning R genes from wild crops is hindered by the requirement for segregating or mutant progenies. Sanu Arora, from the John Innes Centre, and colleagues now report a strategy for rapid R gene cloning by coupling association genetics with R gene enrichment sequencing (AgRenSeq).

To test the AgRenSeq method, the researchers obtained a panel of Aegilops tauschii (diploid wheat) lines that were phenotyped using six races of the wheat stem rust pathogen. A sequence capture bait library was designed and optimized for capturing nucleotide-binding/leucine-rich (NLR) sequences encoded by the R genes in this population. The enriched R gene sequences were then assembled into NLR contigs and NLR k-mers were extracted for each accession. After a pre-filtering step, k-mer-based association mapping was conducted to identify k-mers associated with the resistance trait.

Four stem rust resistance genes, Sr33, Sr45, Sr46 and SrTA1662, against three races of the stem rust pathogen were identified using this approach. The function of these genes in conferring resistance was underpinned by previous map-based or mutagenesis cloning and transgenic data, indicating the power of AgRenSeq. Rapid R gene cloning by AgRenSeq will facilitate marker-assisted breeding and broad-spectrum resistance engineering in genetically modified crops without a need for a reference genome.

However, AgRenSeq will likely miss atypical R genes because it relies on the capture of NLR sequences. Additionally, its performance is affected by sample size and population structure.