WUSCHEL-related homeobox (WOX) proteins participate profoundly in plant development and stress responses. As the difficulty of somatic embryogenesis severely constrains cotton genetic modification, in this study, we identified and comprehensively analyzedWOXgenes in cotton. As a result, 40WOXgenes were identified in the upland cotton genome. All these cottonWOXgenes were classified into three clades, ancient, intermediate, and modern clades, based on the phylogenetic analysis of previous studies. The majority (24) of the cottonWOXgenes belonged to the modern clade, in which all gene members contain the vital functional domain WUS-box, which is necessary for plant stem cell regulation and maintenance. Collinearity analysis indicated that theWOXgene family in cotton expanded to some degree compared to Arabidopsis, especially in the modern clade. Genome duplication and segmental duplication may greatly contribute to expansion. Hormone-response- and abiotic-stress-response-related cis-acting regulatory elements were widely distributed in the promoter regions of cottonWOXgenes, suggesting that the corresponding functions of stress responses and the participation of development processes were involved in hormone responses. By RNA sequencing, we profiled the expression patterns of cottonWOXgenes in somatic embryogenesis. Only about half of cottonWOXgenes were actively expressed during somatic embryogenesis; different cottonWOXgenes may function in different development stages. The most representative,GhWOX4andGhWOX13, may function in almost all stages of somatic embryogenesis;GhWOX2andGhWOX9function in the late stages of embryo patterning and embryo development during cotton somatic embryogenesis. Co-expression analysis showed that the cottonWOXsco-expressed with genes involved in extensive genetic information processing, including DNA replication, DNA repair, homologous recombination, RNA transport, protein processing, and several signaling and metabolism pathways, in which plant hormones signal transduction, MAPK signaling pathways, phosphatidylinositol signaling systems, and ABC transporters, as well as the metabolism of fatty acid; valine, leucine, and isoleucine biosynthesis; and cutin, suberine, and wax biosynthesis, were most significantly enriched. Taken together, the present study provides useful information and new insights into the functions of cottonWOXgenes during somatic embryogenesis. The specific regulatory roles of someWOXgenes in somatic embryogenesis are worthy of further functional research.