Water-soluble humic materials (WSHMs) can enhance the nodule numbers of soybean plants. In this study, targeted metabolomics and transcriptomics were used to understand this mechanism. Results showed that 500 mg/L WSHM increased the adsorption and colonization of rhizobia in soybean roots. High-performance liquid chromatography and targeted metabolomics showed that WSHMs could regulate the content and distribution of endogenous hormones of soybean plants at the initial stage of soybean nodulation. Transcriptomic analysis showed a total of 2406 differentially expressed genes (DEGs) by the 25th day, accounting for 4.89% of total annotation genes (49159). These DEGs were found to contribute primarily to the MAPK signaling pathway, glycolysis/gluconeogenesis, and plant hormone signal transduction according to the −log 10 (Padjust) value in the KEGG pathway. Subsequently, DEGs related to these hormones were selected for verification using quantity-PCR. The WSHM increased the number of nodules by regulating the expression of endogenous hormones in soybean plants.

IntroductionNitrogen fertilizer intake promotes soybean growth before the formation of nodules, but excess nitrogen has an inhibitory effect on soybean nodulation. It is important to balance nitrogen levels to meet both growth and nodulation needs.Methodsthe nitrogen level suitable for soybean growth and nodulation was studied, the role of humic acid (HA) in alleviating the inhibition of high nitrogen on soybean nodulation was analyzed, and transcriptomic analysis was performed to understand its mechanism.ResultsThe results showed that a lower level of nitrogen with 36.4 mg urea per pot could increase the number of nodules of soybean, and a higher level of nitrogen with 145.9 mg urea per pot (U4 group) had the best growth indicators but inhibited nodulation significantly. HA relieved the inhibitory effect at high nitrogen level, and the number of nodules increased by 122.1% when 1.29 g HA was added (H2 group) compared with the U4 group. The transcriptome analysis was subsequently performed on the H2 and U4 groups, showing that there were 2995 differentially expressed genes (DEGs) on the 25th day, accounting for 6.678% of the total annotated genes (44,848) under the test conditions. These DEGs were enriched in mitogen-activated protein kinase signaling pathway-plant, flavonoid biosynthesis, and plant hormone signal transduction based on the –log10 (Padjusted) value in the Kyoto Encyclopedia of Genes and Genomes pathway (KEGG).DiscussionHA balanced the nitrogen level through the above pathways in soybean planting to control the number of nodules.