TRANSFORMATION OF ORNAMENTAL TOBACCO AND KALE MEDIATED BY AGROBACTERIUM-TUMEFACIENS AND A-RHIZOGENES HARBORING A REPORTER, BETA-GLUCURONIDASE (GUS) GENE

Transformation of ornamental tobacco (Nicotiana X Sanderae Hort.) and kale (Brassica oleracea var. acephala DC) was studied using the Agroba cterium tumefaciens or A. rhizogenes-vector system. A plasmid, pBI 121 (beta-glucuronidase (GUS) gene combined with kanamycin resistant gene , nptII) was transferred to Agrobacterium by tri-parental mating or fr eeze-thaw method. After leaf disks of ornamental disks were inoculated with a suspension of A. tumefaciens harboring pBI 121, they were cult ured on Murashige and Skoog (MS) medium supplemented with growth regul ators, kanamycin sulfate and carbenicillin sodium. Six calli appeared from edges of leaf disks from which shoots differentiated. Of the 10 s hoots examined, nine showed positive reaction to 5 bromo-4-chloro-3-in dolyl glucuronide (X-gluc) suggesting that GuS gene was transferred to plant cells. These nine shoots survived on MS medium containing kanam ycin sulfate whereas shoots regenerated from non-infected leaf disks d ied. Based on these results, nine shoots were recognized to be transfo rmants harboring pBI 121. Petioles of ornamental kale leaves were inoc ulated with a suspension of A. rhizogenes harboring a plasmid, pBI 121 . Ten roots developed, but only two roots showed positive reaction to X-gluc. Five shoots which were regenerated from these two roots on MS medium supplemented with growth regulators and carbenicillin sodium ga ve a positive reaction to X-gluc, and to 4-methyl umbelliferyl glucuro nide (MUG). The shoots survived on MS medium containing kanamycin sulf ate, indicating that the five regenerated shoots were transformed.