Characterization and Fine Mapping of the Stay-Green-Related Spot Leaf Gene TaSpl1 with Enhanced Stripe Rust and Powdery Mildew Resistance in Wheat

Lesion mimic phenotypes, characterized by leaf spots formed in the absence of pathogens or pests, are often associated with reactive oxygen species (ROS) accumulation and cell necrosis. This study identified a novel and stable homozygous spotted phenotype (HSP) from the F₈ population of common wheat (XN509 × N07216). The yellow spots that appeared at the booting stage were light-sensitive, and accompanied by cell necrosis and H₂O₂ accumulation. Compared with homozygous normal plants (HNPs), HSPs exhibited enhanced resistance to stripe rust and powdery mildew without compromising yield. RNA-Seq analysis at three stages revealed that differentially expressed genes (DEGs) between HSPs and HNPs were significantly enriched in KEGG pathways related to photosynthesis and photosynthesis-antenna proteins. GO analysis highlighted chloroplast and light stimulus-related down-regulated DEGs. Fine mapping identified <i>TaSpl1</i> within a 0.91 Mb interval on chromosome 3DS, flanked by the markers <i>KASP188</i> and <i>KASP229</i>, using two segregating populations comprising 1117 individuals. The candidate region contained 42 annotated genes, including 14 DEGs based on previous BSR-Seq data. PCR amplification and qRT-PCR verification identified the expression of <i>TraesCS3D02G022100</i> was consistent with RNA-Seq data. Gene homology analysis and silencing experiments confirmed that <i>TraesCS3D02G022100</i> was associated with stay-green traits. These findings provide new insights into the genetic regulation of lesion mimics, photosynthesis, and disease resistance in wheat.