Rapeseed (Brassica napus L.) is a major oil crop worldwide that is vigorously promoted for cultivation in China. Boron (B) is an essential micronutrient for plant growth and development. However, agricultural soils in rapeseed planting areas often show either B deficiency or severe B deficiency. Increasing the resistance to B deficiency is a pivotal goal in the breeding of rapeseed, yet the genetic basis for variations in B efficiency-related traits remains unclear. In this study, a natural population with 391 rapeseed accessions was used to investigate B efficiency-related traits through a nutrient solution system, including relative root length (RRL), shoot dry weight (SDW), root dry weight (RDW), and B efficiency coefficient (BEC), which exhibited extensive phenotypic variations under B deficiency. Through a genome-wide association study (GWAS) of B efficiency-related traits using high-density SNP markers obtained from whole-genome resequencing, we identified 106 significantly associated SNPs by employing both the general linear model and the mixed linear model. Among these SNP loci, two prominent SNP clusters were detected on chrA03:14087835–14764672 and chrC03:20110319–22135492 at low B level across three repeated experiments of multiple traits. Integrated with the transcriptome analysis, four genes, BnaA03g29020D, BnaA03g29440D, BnaC03g33010D, and BnaC03g34490D, exhibiting higher differentially expressed fold-change along with favorable haplotypes within the promoter or coding region were identified as candidate genes that could potentially be involved in B efficient utilization, and their favorable haplotypes improved seedling growth and productivity under B deficiency. In view of the lack of B mineral resources in China, rapid and accurate identification of more B-efficient alleles and the study of the genetic mechanism underlying crops in response to B deficiency have important theoretical and practical significance for cultivating B-efficient varieties and maintaining green, sustainable agriculture quickly and accurately.1
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