Introduction Lentil ( Lens culinaris Medik .) is a nutrient-dense legume that plays a critical role in global food security and human health. In addition to being a rich source of protein, fiber, and micronutrients, lentil contains resistant starch (RS), a component of dietary fiber associated with improved glycemic response, enhanced gut microbiota, and reduced inflammation. However, the genetic basis of RS variation in lentil remains poorly understood. Methods Resistant starch concentration, total starch concentration, 100-seed weight, and plot seed weight were quantified in a panel of 192 diverse lentil accessions and five checks evaluated across three environments. Genome-wide association studies (GWAS) were conducted using 5,583 high-quality single nucleotide polymorphisms (SNPs). Putative candidate genes were identified within 100-kb windows of significant marker -trait associations (MTAs), followed by gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. Results Nine significant MTAs were identified for RS, three for total starch, 20 for 100-seed weight, and 11 for plot seed weight, explaining 2.9%–42.6% of the phenotypic variance. Putative candidate genes included soluble starch synthase III (SS3), which is involved in starch biosynthesis. Functional enrichment analyses revealed overrepresentation of biological processes associated with oxidative stress response, detoxification, starch and amylopectin biosynthesis, ATP synthesis, and auxin transport. Enriched molecular functions included peroxidase, peptidyl -prolyl isomerase, and UDP-glucosyltransferase activities, while enriched cellular components included the chloroplast, peroxisome, vacuole, and cell junctions. Discussion These findings provide insights into the genetic basis of resistant starch accumulation in lentil seeds and identify potential genetic targets for future breeding efforts aimed at improving the nutritional quality of lentil cultivars.
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