The CO family genes regulate flowering time by integrating circadian rhythms, light signals, and meristem-related genes through the photoperiod pathway, and play a crucial role in the process of plant flower formation. In this study, a CO family gene cloned from Chrysanthemum lavandulifolium by RT-PCR method, was named ClCOL16 (GenBank accession number: AOA52174.1). Arabidopsis thaliana was transformed by inflorescence immersion mediated by Agrobacterium tumefaciens, and T3 generation transgenic plants were selected by resistant screening and PCR detection of the heterologous COL gene. To analysis the function of ClCOL16 in plant photoperiod regulation of flowering and lay the foundation for the further study for flowering regulation mechanism in Chrysanthemum morifolium, we cultured six transgenic lines randomly selected and the wild type (WT) of A. thaliana in a long day environment (alternating light and dark at 16 h/ 8 h) to observe their phenotypes, and detected the expression of ClCOL16 in WT and transgenic A. thaliana leaves after the 37 days of planting. The results showed: (1) ClCOL16 gene of C. lavandulifolium ORF was 1 278 bp and encoding a protein of 425 amino acids; Structural analysis showed ClCOL16 protein contains a B-box and a CCT conserved domain, which is a typical characteristic of CO family Group Ⅲ. (2) NCBI blast analysis showed that ClCOL16 protein had the highest identity with COL16 (GenBank accession number: GEZ38716.1) from Tanacetum cinerariifolium, up to 94.38%, which further proved that it belongs to the CO family Group Ⅲ. (3) Expression analysis showed that ClCOL16 gene was constitutive expression in C. lavandulifolium, and the highest expression was in leaves； ClCOL16 gene was regulated by circadian clock, under short-day condition its expression showing obvious circadian rhythm. (4) Ectopic expression showed that compared to wild type A. thaliana, the flowering time of ClCOL16 transgenic lines was advanced by 7 days, the number of rosette leaves was reduced by 3 to 4, and the expression of ClCOL16 gene in leaves was significantly increased. All the results indicated that ClCOL16 gene ectopic expression lead to early flowering in A. thaliana. According to the experimental results, we speculated that ClCOL16 gene may be an important gene involved in responding to light signal, and participating in the regulation of photoperiodic flowering pathway in C. morifolium.