Abstract: In order to explore the key genes in response to early symbiotic nitrogen fixation in soybean roots as well as the functions and metabolic pathways of these key genes, this study used transcriptomics technology to perform RNA sequencing on soybean root materials one day after with or without inoculation of rhizobia. The key genes involved in the early symbiotic nitrogen fixation process were explored by GO enrichment and KEGG enrichment analysis of differentially expressed genes, and the transcription factors regulated by rhizobia were further screened. The AP2/ERF family members with the largest number of enrichments were subjected to evolutionary analysis. Finally, the expression of some genes was verified by qRT-PCR. A total of 824 differentially expressed genes (DEGs) in response to rhizobium infection are identified. GO enrichment and KEGG enrichment analysis showed that DEGs are mainly enriched in transcriptional regulation, hormone-related and secondary metabolite synthesis pathways. Phylogenetic analysis showed that AP2/ERF family members in soybean could be clustered into three branches according to their genetic relationship with MtERN1/2 and LjERN. Among them, branch I may play a role in inducing rhizobium infection and nodule organogenesis. The results of qRT-PCR showed that the AP2/ERF transcription factor is indeed regulated to varying degrees after inoculation with rhizobia. The DEGs found in this study provide candidate molecules for a better understanding of the early symbiotic nitrogen fixation mechanism.