In the mammalian tooth development system, tooth morphogenesis is strictly regulated by complex genetic networks that govern the formation of different tooth shapes and types. Previous studies about tooth development mainly focused on mandibular teeth; the gene expression pattern of maxillary tooth development and its differences remain largely unknown. RNA sequencing (RNA-seq) analysis was employed to obtain gene expression profiles of the maxillary and mandibular tooth germs at the cap stage. Differential gene expression was assessed using gene ontology enrichment analysis and protein-protein interaction network analysis. The significant differential gene expression data between the maxillary and mandibular tooth germs were validated using quantitative real-time PCR (RT-qPCR) and immunofluorescence (IF). The transcriptome expression profiles of the maxillary and mandibular tooth germs at the cap stage in human fetuses were successfully constructed. Transcriptome analysis revealed shared activation of the TGF-β, WNT, and NOTCH pathways, with the maxillary tooth germs biased toward morphogenesis and signaling and the mandibular tooth germs towards mineralization and metabolism. A total of 687 differentially expressed genes were identified, highlighting regional differences. RT-qPCR and IF validation confirmed the expression of the key genes. Protein–protein interaction analysis identified core regulators, suggesting their pivotal roles in region-specific tooth development. These findings suggest a region-specific regulatory pattern in human cap-stage tooth germ development, with a tendency toward morphological specialization in the maxillary tooth germs, and mineralization-related processes in the mandibular tooth germs.