The three-domain multicopper oxidase (TMCO) family, the largest subfamily of multicopper oxidases, plays a crucial role in plant growth, development, and responses to biotic stresses. To investigate the expression characteristics of the SbTMCO gene family under aphid infestation, a genome-wide identification and analysis of TMCO family members in sorghum were conducted using bioinformatics methods. Transcriptome data and qRT-PCR were utilized to preliminarily examine sorghums responses to aphid stress. The results revealed 42 TMCO genes in the sorghum genome, distributed across 10 chromosomes and classified into five subgroups. Members within the same subgroup displayed similar physicochemical properties, gene structures, and conserved motifs. Collinearity analysis suggested that tandem duplication and segmental duplication were the primary drivers of TMCO gene expansion in sorghum. Cis-acting element analysis identified multiple jasmonic acid (MeJA) and abscisic acid (ABA) responsive elements among SbTMCO genes. Gene expression analysis, based on RNA-seq and qRT-PCR, confirmed the relative expression levels of TMCO genes under aphid stress. This study establishes a foundation for exploring the anti-aphid functions of SbTMCO genes and provides recommendations and directions for further research on this gene family.