Overexpression of Doritaenopsis Hybrid EARLY FLOWERING 4-like4 Gene, DhEFL4, Postpones Flowering in Transgenic Arabidopsis

The EARLY FLOWERING 4 (ELF4) gene has been reported as a requirement for photoperiod perception and circadian function. In this study, an ELF4 homolog gene EARLY FLOWERING 4-like4 (EFL4) was isolated from Doritaenopsis “Tinny Tender” (Doritaenopsis Happy Smile × Happy Valentine) and designated as DhEFL4. The acquired DhEFL4 cDNA was 593 bp with a 345 bp open reading frame encoding 114 amino acids. The putative DhEFL4 protein contained a conserved region, DUF1313, which is shared by the ELF4 family. Amino acid sequence alignment showed that DhEFL4 shared high similarity to ELF4 and ELF4-like homologs from other species. The DhEFL4 promoter was also obtained, and 241 cis-elements were found. The diurnal expression of DhEFL4, which may be affected by the EVENINGAT (EE) and circadian motif was analyzed by quantitative PCR under short- and long-day conditions. Reverse-transcription PCR analysis results showed that DhEFL4 is an evening-expressed circadian gene. DhEFL4 was expressed in both vegetative and reproductive organs, and its transcripts reached higher levels in the buds. To further study the function of the DhEFL4 gene, DhEFL4 was overexpressed in Arabidopsis thaliana by Agrobacterium-mediated transformation. The flowering of these transgenic plants was more delayed than that of wild-type Arabidopsis by 12−14 days, and flower stems increased. Results indicated that DhEFL4 may perform an important function in the regulation of flowering.