In this study, a flavanone 3hydroxylase (F3H) gene named CmF3H was cloned from Chrysanthemum morifolium ‘14C1’. Bioinformatics analysis showed that the full length of ‘14C1’ CmF3H cDNA (GenBank: MW454869) is 1 284 bp, and the open reading frame length is 1 095 bp, encoding 364 amino acids. The theoretical molecular weight of ‘14C1’ CmF3H coding protein is 41.19 kD, theoretical pI is 5.57, the instability index is 39.51, the grand average hydropathicity is -0.465, the aliphatic index is 83.02. Amino acid alignment results indicated that ‘14C1’ CmF3H protein belongs to 2oxoglutaratedependent dioxygenase (2ODD) protein family, and has the typical characteristics of 2ODD conservative structure domain. Phylogenetic tree analysis showed that ‘14C1’ and C. morifolium ‘SU07’ were located on the same evolutionary node, which indicated that they had the closest relationship. The promoter of ‘14C1’ CmF3H was cloned by Genome Walking Method (GenBank: MW463894), its full length is 1 217 bp. The promoter sequence included light responsive element, drought and ABA responsive elements, MYB recognition and binding site as well as light responsive and organization specific expression elements. The analysis of realtime quantitative PCR indicated that CmF3H gene was expressed among different tissues of ‘14C1’ such as root, stem, leaf, bud, ligulate flower and tubular flower, the expression levels of CmF3H were the highest in tubular flower, followed by stem, leaf, bud, ligulate flower, and lowest in root. The results of this study can provide reference for the study on the biosynthesis mechanism of flavonoids in C. morifolium.