The cytochrome P450 monooxygenases (CYP450) are the largest enzyme family in plant metabolism and widely involved in the biosynthesis of primary and secondary metabolites. Foxtail millet (Setaria italica(L.) P.Beauv) can respond to abiotic stress through a highly complex polygene regulatory network, in which the SiCYP450 family is also involved. Although the CYP450 superfamily has been systematically studied in a few species, the research on the CYP450 superfamily in foxtail millet has not been completed. In this study, three hundred and thirty-oneSiCYP450genes were identified in the foxtail millet genome by bioinformatics methods, which were divided into four groups, including forty-six subgroups. One hundred and sixteen genes were distributed in thirty-three tandem duplicated gene clusters. Chromosome mapping showed that SiCYP450 was distributed on seven chromosomes. In the SiCYP450 family of foxtail millet, 20 conserved motifs were identified.Cis-acting elements in the promoter region ofSiCYP450genes showed that hormone response elements were found in allSiCYP450genes. Of the three hundred and thirty-oneSiCYP450genes, nine genes were colinear with theArabidopsis thalianagenes. Two hundredSiCYP450genes were colinear with theSetaria viridisgenes, including two hundred and forty-five gene duplication events. The expression profiles ofSiCYP450genes in different organs and developmental stages showed thatSiCYP450was preferentially expressed in specific tissues, and many tissue-specific genes were identified, such asSiCYP75B6,SiCYP96A7,SiCYP71A55,SiCYP71A61, andSiCYP71A62in the root,SiCYP78A1andSiCYP94D9in leaves, andSiCYP78A6in the ear. The RT-PCR data showed thatSiCYP450could respond to abiotic stresses, ABA, and herbicides in foxtail millet. Among them, the expression levels ofSiCYP709B4,SiCYP71A11,SiCYP71A14,SiCYP78A1,SiCYP94C3, andSiCYP94C4were significantly increased under the treatment of mesotrione, florasulam, nicosulfuron, fluroxypyr, and sethoxydim, indicating that the same gene might respond to multiple herbicides. The results of this study will help reveal the biological functions of the SiCYP450 family in development regulation and stress response and provide a basis for molecular breeding of foxtail millet.