The Brassica napus extA promoter: a novel alternative promoter to CaMV 35Sfor directing transgene expression to young stem tissues and load bearing regions of transgenic apple trees (Malus pumila Mill.)

The Brassica napus extensin A gene is highly expressed in root tissue of oi lseed rape. In an attempt to identify an effective root-specific promoter f or biotechnological applications, we have examined the ability of the -940 extA promoter to drive expression of the gusA reporter gene in the vegetati ve tissues of apple (Malus pumila Mill cv Greensleeves). Transgenic apple l ines were produced by Agrobacterium tumefaciens-mediated transforation and GUS activity was analysed both quantitatively and qualitatively. The extA p romoter was active in all tissues of young plants in all 15 clones examined . However Southern blot data suggested that only a proportion of the popula tion contained the entire promoter and that others had suffered deletion of unknown length. This may have contributed to the variation seen in the qua ntitative and qualitative expression of GUS. Specific GUS activity was high est in the stems where it approached, an din some clones, exceeded that usi ng the constitutive CaMV 35S promoter. Histochemical analysis confirmed tha t GUS was localised to tissues involved in structural support of the stem. Staining was particularly intense at nodal junctions where high tensile str ess is exerted on the tissues. maturing phloem tissues showed localisation of expression to the phloem parenchyma cells and phloem fibres. Transverse sections of the root revealed staining of primary procambial tissues includ ing the young endodermis but not staining was seen in the cortex. Although the -940 extA promoter is clearly not root-specific in apple, it is likely to have useful biotechnological applications in tree species.