Functional Characterization of GmRD22 Modulating Nitrogenase Activity in Soybean with Transcriptomic Comparison Between Two Genotypes

Soybean (Glycine max) is a key crop in China grown as a source of edible oil and plant-derived protein, and its production is closely linked to national food security. Insufficient nitrogen availability remains a key constraint on soybean yield. In legumes, symbiotic nitrogen fixation (SNF) enables the conversion of atmospheric nitrogen into bioavailable forms, thereby reducing reliance on synthetic fertilizers and improving soil quality. Nitrogenase activity is a central determinant of SNF efficiency; however, its regulatory mechanisms in soybean nodules are not yet fully understood. In this study, transcriptomic data from two soybean accessions with contrasting SNF performance (Suinong 14 and ZYD00006) were analyzed, leading to the identification of Glyma.04G013500 as a member of the GmRD22 gene family. This study verified that this gene is potentially associated with nitrogenase activity. Functional characterization revealed that this gene acts as a negative regulator of nodulation by influencing the expression of genes associated with nodule development. Haplotype analysis further uncovered a pattern consistent with domestication, as the elite haplotype (HapI) with enhanced nitrogen fixation capacity, exhibited a progressive increase in frequency from wild soybean populations to landraces and modern cultivars. These findings suggest that GmRD22 has undergone directional selection during soybean domestication and improvement. Overall, these results offer new insights into the genetic control of SNF and establish promising targets for breeding soybean varieties with improved nitrogen fixation efficiency.