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Excised leaf sections of lance coreopsis cultured on Murashige Skoog (MS) medium produced adventitious shoots in response to BA. When the combinations of 0, 0.5, 1, or 2 μm NAA with 0, 5, 10, 20, or 40 μm BA were tested, shoots were induced by any of the four BA concentrations used in the medium, regardless of the presence of NAA. The average number of shoots formed per leaf section ranged from 1.4 to 4.3 seven weeks after culture initiation. Roots were induced at the base of individual shoots on the same regeneration medium when cultures were kept longer than 7 weeks. The rooted plants were transferred successfully into soil. The regenerated plants had the same growth and flowering characteristics as the seed-grown plants. Chemical names used: benzyladenine (BA); naphthaleneacetic acid (NAA).
Leaf discs of Coreopsis lanceolata, when cultured on Murashige and Skoog (MS) medium supplemented with 10 μM benzyladenine (BA) and 1 μM naphthaleneacetic acid (NAA), produced shoots in four weeks. Shoots were often induced on the marginal tissues of the leaf discs without callus formation. The frequency and the number of shoots induced per leaf disc varied slightly when growth regulator combinations of 0.5-40 μM BA and 0-2 μM NAA were tested. Most shoots produced roots on the same regeneration medium after formation of 3 to 5 leaves. The rooted plants were established in soil within 3 months from initial culture. For genetic transformation, leaf discs were infected with Agrobacterium tumefaciens Strain LBA 4404 carrying both kanamycin resistance and β-glucuronidase (GUS) genes. Shoots were induced from these leaf discs on the regeneration medium containing 250 mg/liter kanamycin. The histochemical assay showed that the regenerated shoots were GUS positive.
Leaf discs of Salpiglossis sinuata L. were infected with disarmed Agrobacterium tumefaciens strain LBA 4404 carrying β-glucuronidase (GUS) gene and kanamycin resistance. Plantlets were regenerated from the leaf discs cultured on Murashige and Skoog (MS) medium supplemented with 10 μM kinetin and 1 μM naphthaleneacetic acid (NAA) plus 300 mg/liter kanamycin. The plantlets were then rooted on the MS medium containing the same concentration of kanamycin plus 2 μM NAA without kinetin. The histochemical assay showed that the regenerated plants were GUS positive. A Southern blot test is underway to determine the stability of the transferred genes in the regenerated plants.
The phenotypic expression and inheritance of the rolC gene in the transgenic plants of Salpiglossis sinuata L. were investigated. The chasmogamous salpiglossis plants with solid yellow flower color (ccrrDD) were transformed with Agrobacterium tumefaciens strains LBA4404 and EHA101 carrying rolC, GUS, and NPTII genes via a leaf disc co-cultivation system. The transgenic plants were shorter in plant height, produced more branches with a compact growth habit, and developed smaller flowers and narrower leaves as compared to the control plant. While the transgenic plants showed the same corolla color and color shades as the parental line, they became male sterile. A backcross between a male-sterile transgenic plant (ccrrDD plus rolC) and a nontransformed red-flowering line (ccRRDD) produced a progeny with red flower color and the same altered growth habit as the transgenic female parent. Only 4 out of 32 plants in this progeny population showed the negative GUS staining as well as the non transgenic phenotype. These results suggest that at least two copies of the rolC gene were integrated into one homologous chromosome pair during transformation and that a cross-over event may have occurred during meiosis.