Large-scale identification and functional analysis of <i>NLR</i> genes 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 more <i>NLR</i> genes than other cultivars? Or does Tetep possess multiple major <i>NLR</i> genes that can individually confer broad-spectrum resistance to blast? Moreover, are there many interacting <i>NLR</i> pairs in the Tetep genome? We sequenced its genome, obtained a high-quality assembly, and annotated 455 nucleotide-binding site leucine-rich repeat (<i>NLR</i>) genes. We cloned and tested 219 <i>NLR</i> genes 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 cloned <i>NLR</i>s showed resistance to 1 or more pathogen strains and each strain was recognized by multiple <i>NLR</i>s. However, few <i>NLR</i>s showed resistance to >6 strains, so multiple <i>NLR</i>s 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-derived <i>NLR</i>s. In developing a method to identify <i>NLR</i> pairs each of which functions as a unit, we found that >20% of the <i>NLR</i>s in the Tetep and 3 other rice genomes are paired. Finally, we designed an extensive set of molecular markers for rapidly introducing clustered and paired <i>NLR</i>s 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.