Abstract
SummaryPlant resistance (R) and pathogen avirulence (Avr) gene interactions play a vital role in pathogen resistance. Efficient molecular screening tools for crops lack far behind their model organism counterparts, yet they are essential to rapidly identify agriculturally important molecular interactions that trigger host resistance.Here, we have developed a novel wheat protoplast assay that enables efficient screening of Avr/R interactions at scale. Our assay allows access to the extensive gene pool of phenotypically describedRgenes because it does not require the overexpression of clonedRgenes. It is suitable for multiplexedAvrscreening, with interactions tested in pools of up to fiftyAvrcandidates.We identified Avr/R-induced defense genes to create promoter-luciferase reporter. Then, we combined this with a dual-color ratiometric reporter system that normalizes read-outs accounting for experimental variability and Avr/R-induced cell-death. Moreover, we introduced a self-replicative plasmid reducing the amount of plasmid used in the assay.Our assay increases the throughput ofAvrcandidate screening, accelerating the study of cellular defense signaling and resistance gene identification in wheat. We anticipate that our assay will significantly accelerateAvridentification for many wheat pathogens, leading to improved genome-guided pathogen surveillance and breeding of disease-resistant crops.
Publisher
Cold Spring Harbor Laboratory
Cited by
3 articles.
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