Large-scale identification and functional analysis ofNLRgenes in blast resistance in the Tetep rice genome sequence

Abstract

Tetep is a rice cultivar known for broad-spectrum resistance to blast, a devastating fungal disease. The molecular basis for its broad-spectrum resistance is still poorly understood. Is it because Tetep has many moreNLRgenes than other cultivars? Or does Tetep possess multiple majorNLRgenes that can individually confer broad-spectrum resistance to blast? Moreover, are there many interactingNLRpairs in the Tetep genome? We sequenced its genome, obtained a high-quality assembly, and annotated 455 nucleotide-binding site leucine-rich repeat (NLR) genes. We cloned and tested 219NLRgenes as transgenes in 2 susceptible cultivars using 5 to 12 diversified pathogen strains; in many cases, fewer than 12 strains were successfully cultured for testing. Ninety clonedNLRs showed resistance to 1 or more pathogen strains and each strain was recognized by multipleNLRs. However, fewNLRs showed resistance to >6 strains, so multipleNLRs are apparently required for Tetep’s broad-spectrum resistance to blast. This was further supported by the pedigree analyses, which suggested a correlation between resistance and the number of Tetep-derivedNLRs. In developing a method to identifyNLRpairs each of which functions as a unit, we found that >20% of theNLRs in the Tetep and 3 other rice genomes are paired. Finally, we designed an extensive set of molecular markers for rapidly introducing clustered and pairedNLRs in the Tetep genome for breeding new resistant cultivars. This study increased our understanding of the genetic basis of broad-spectrum blast resistance in rice.