Isolation of an actin promoter for strong expression of transgenes in the orchid genus Dendrobium

We isolated and functionally characterized a Dendrobium Actinl (DmACT1) promoter that drives strong gene expression in the orchid genus Dendrobium. A genomic fragment containing the region 3227 bp upstream of the coding region of DmACT1 was obtained by inverse PCR. Detailed comparison of the full-length cDNA and genomic sequences revealed that DmACT1 has a 1374 bp first intron in the 5`` UTR. However, the 5`` flanking sequences upstream of the coding region showed no obvious sequence similarities compared to those of known promoters, including plant actin promoters. Serial deletion constructs of the 5`` flanking region from the translation initiation codon were fused to the coding sequence of a GUS/Iuciferase fusion reporter to identify the regulatory element necessary for promoter activity. Transient assays in the flowers of Dendrobium revealed that the 5`` UTR-intron greatly enhanced pro-moter activity. Moreover, the DmACT1 promoter with its 5`` UTR-intron yielded approximately 10-fold higher reporter activity than the rice Actl promoter-intron. Our data suggest that the DmACT1 promoter with its 5`` UTR-intron is a useful tool for strong expression of transgenes in Dendrobium orchids.