Comparative and Evolutionary Analysis of Single-Copy Genes in Brassica napus Identifies BnaNRA1 as a Contributor to Nutrient-Responsive Lateral Root Development

Single-copy genes are important components of plant genomes, but their evolutionary origins and biological functions in polyploid crops remain poorly understood. The objective of this study was to clarify the evolutionary origins of single-copy genes in allotetraploid Brassica napus and identify nutrient-responsive single-copy genes involved in root development. Here, we systematically identified single-copy genes across 16 representative plant species and investigated their evolution and potential functions in allotetraploid Brassica napus. A total of 167,219 single-copy genes were identified across the 16 species, including 5106 genes in the B. napus cultivar Zhongshuang 11 (ZS11). Comparative synteny analysis with B. rapa and B. oleracea classified 204 chromosome-anchored genes with unambiguous parental relationships into 38 inherited and 166 inferred loss-derived single-copy genes, indicating that post-polyploid duplicate loss is an important route for the re-establishment of single-copy states in B. napus. Integration of promoter and expression analyses identified 65 root- and nutrient-responsive candidates. Among them, BnaA09G0594900ZS, designated BnaNRA1, was induced by low-nitrogen and low-phosphorus treatments and encoded a predominantly nuclear-localized protein. Three independent bnanra1-edited lines showed no consistent root phenotype under nutrient-sufficient conditions but exhibited reduced lateral root formation under nitrogen and phosphorus deficiency, with a particularly consistent effect under phosphorus deficiency. Collectively, these findings indicate that post-polyploid duplicate loss contributes to the re-establishment of single-copy states in B. napus and identify BnaNRA1 as a nutrient-responsive single-copy gene contributing to lateral root development under nutrient limitation.