Rapid identification disease resistance genes from plant genomes by resistance gene enrichment sequencing (RenSeq) of EMS-derived susceptible mutants

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Wild relatives of domesticated crops contain many useful disease resistance (R) genes. Introducing this natural resistance is an elegant way of managing disease. However, traditional methods for introducing R genes typically involve long breeding trajectories to avoid linkage drag, i.e. the simultaneous introduction of deleterious traits. Furthermore, R genes tend to be overcome by the pathogen within a few seasons when deployed one at a time.

Our long-term strategy is to isolate, by molecular cloning, as many new R genes as possible, and introduce them in combinations using GM methods. Molecular cloning makes it possible to put several new genes together in the same location in the genome, allowing breeders to work with them as a “single” gene and avoiding linkage drag. Moreover, a pyramid of R genes with distinct specificities should be more durable.

Traditional map-based cloning of R genes, however, is still challenging. First, large tracts of plant genomes are inaccessible to map-based genetics due to low recombination. Second, most R genes belong to a structural class of genes called NB-LRRs, which tend to reside in complex clusters, and many hundreds of NB-LRRs populate a typical plant genome.

We propose a strategy that will significantly increase the rate of R gene identification. In a first step, we will screen large numbers of EMS mutagenized plants for susceptible mutants. Secondly, we will use a state-of-the art sequencing technique recently implemented in our lab to selectively capture and sequence all the NB-LRRs in a plant genome. This will allow rapid and cheap comparison of wildtype with mutants to identify and clone R genes. The outputs of this research will be three-fold: (i) using known controls we will implement our generic strategy to isolate R genes from complex genomes, (ii) we will apply this strategy to identify novel R genes from potato and wheat, and (iii) we will test our key wheat rust R genes in Indian and UK environments.

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