We investigated the effects of maternal vitamin and mineral supplementation throughout gestation on gene expression in the jejunal mucosa of neonatal calves. Crossbred Angus heifers (n = 14) were estrus synchronized, bred to female-sexed semen, and randomly assigned to a basal diet (Control, CON; n = 7) or the basal diet plus vitamin and mineral supplement (Treatment, VTM; n = 7). After parturition, calves were removed from their dams before suckling, fed colostrum replacer, and euthanized 30 h after the first feeding. A subsample of the mucosa of the mid-jejunum was collected, and total RNA was isolated. Gene expression was measured using RNA-Seq, and differentially expressed genes (DEGs) were identified using DESeq2. We identified 528 DEGs from the jejunal mucosa between the VTM and CON calves (P ≤ 0.05 and |log2FC| ≥ 0.5). The DEGs were associated with nutrient transport, lipid metabolism, and immune-related biological processes and pathways. Interestingly, genes underlying the complement and coagulation cascades were mostly downregulated in calves born to VTM dams. On the other hand, the cytokine-cytokine receptor interaction KEGG pathway showed most genes upregulated (LIFR, KDR, TNFRSF4, TNFSF18, FLT1, and TNFRSF12A). Our results show that vitamin and mineral supplementation throughout gestation affects genes underlying tissue structure, nutrient transport and metabolism, and immune system pathways in neonates. The implications of such changes and the long-term outcomes on herd health and performance warrant further research.