*Article* **Deciphering the Genetic Basis of Root and Biomass Traits in Rapeseed (***Brassica napus* **L.) through the Integration of GWAS and RNA-Seq under Nitrogen Stress**

**Nazir Ahmad 1,† , Bin Su 1,†, Sani Ibrahim 1,2, Lieqiong Kuang <sup>1</sup> , Ze Tian <sup>1</sup> , Xinfa Wang 1,3, Hanzhong Wang 1,3,\* and Xiaoling Dun 1,\***


**Abstract:** An excellent root system is responsible for crops with high nitrogen-use efficiency (NUE). The current study evaluated the natural variations in 13 root- and biomass-related traits under a low nitrogen (LN) treatment in a rapeseed association panel. The studied traits exhibited significant phenotypic differences with heritabilities ranging from 0.53 to 0.66, and most of the traits showed significant correlations with each other. The genome-wide association study (GWAS) found 51 significant and 30 suggestive trait–SNP associations that integrated into 14 valid quantitative trait loci (QTL) clusters and explained 5.7–21.2% phenotypic variance. In addition, RNA sequencing was performed at two time points to examine the differential expression of genes (DEGs) between high and low NUE lines. In total, 245, 540, and 399 DEGs were identified as LN stress-specific, high nitrogen (HN) condition-specific, and HNLN common DEGs, respectively. An integrated analysis of GWAS, weighted gene co-expression network, and DEGs revealed 16 genes involved in rapeseed root development under LN stress. Previous studies have reported that the homologs of seven out of sixteen potential genes control root growth and NUE. These findings revealed the genetic basis underlying nitrogen stress and provided worthwhile SNPs/genes information for the genetic improvement of NUE in rapeseed.

**Keywords:** rapeseed; root and biomass traits; nitrogen stress; GWAS; RNA sequencing
