Agrobacterium-mediated transformation of the commercially important grapefruit cultivar Rio Red (Citrus paradisi Macf.)

Transgenic plants of grapefruit cv. Rio Red (Citrus paradisi Macf.) have be en obtained by Agrobacterium tumefaciens-mediated gene transfer using seedl ing-derived epicotyl segments as explants and kanamycin as the selective ag ent. The transformation procedure includes a shoot elongation phase with a liquid medium overlay, which provides additional selection against non-tran sgenic shoots. Transformed shoots are invigorated and multiplied on a non-s elective medium prior to grafting, thus assuring that plants can be recover ed from transgenic shoots. We have constructed a binary vector, pBin34SGUS, with an intron-containing P-glucuronidase gene (uidA) under the control of the Figwort mosaic virus 34S promoter. The 34S promoter efficiently drives uidA gene expression both in transient assays and in transgenic Rio Red le af tissue, although at levels five- to sevenfold lower than the Cauliflower mosaic virus 35S promoter. An untranslatable coat protein gene (uncp) of t he Citrus tristeza virus strain SY568 and the Galanthus nivalis agglutinin gene (gna) were inserted into pBin34SGUS and transgenic plants have been ob tained. Stable integration of the uncp and gna genes was confirmed by South ern hybridization and gna gene expression was confirmed by Western blot ana lysis.