Structure of the genomic DNA encoding cucumber ascorbate oxidase and its expression in transgenic plants

Genomic DNA (Aso1) encoding the ascorbate oxidase (ASO) of cucumber (Cucumis sativas) was cloned and sequenced. From a comparison of the sequences of Aso1 and its cDNA [Ohkawa et al. Proc. Natl. Acad. Sci. USA 86 (1989) 1239-1243], it was concluded that Aso1 consisted of four exons and three introns. In the 5'-flanking region, sequence motif resembling a TATA box and a CAAT box, were found 124 and 218 bp upstream, respectively, from the start codon. Sites of initiation of transcription, determined by primer extension, were the A and G residues located 90 and 93 bp, respectively, upstream from the initiation codon. ASO was abundant in cucumber fruits and stems. The 5'-flanking region, placed upstream of the structural gene of β-glucuronidase (GUS), expressed the GUS activity in cucumber plants in a transient expression assay after introduction of DNA with a particle gun. High-level GUS expression was found in stems of transgenic tobacco plants harboring Aso1 promoter-GUS chimeric gene.