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- Author or Editor: E. D. Earle x
Regeneration from VFNT Cherry tomato was optimized prior to transformation. Cotyledon and hypocotyl sections from 7-day-old in vitro grown VFNT Cherry tomato were cultured on medium containing MS salts, B5 vitamins and the following per liter: sucrose, 30g; zeatin, 1 mg; IAA, 0.1 mg; agar, 8.0g or GelriteR, 2.2g. Culture conditions investigated included cotyledon and upper hypocotyl explant lengths, light vs. dark germination, and agar vs. GelriteR. The conditions which resulted in the highest average number of shoots per explant were cotyledon basal explants 2 mm in length, 3.95; cotyledons from dark germination, 6.2; and hypocotyls from light germination, 8.6. An equal number of shoots regenerated on medium containing agar or GelriteR, however, shoots regenerated on medium containing agar were more vigorous. Cotyledon and hypocotyl sections were cocultivated with the Agrobacterium tumefaciens binary vector pBI121 containing the neomycin phosphotransferase II (NPTII) and B-glucuronidasc (GUS) genes. Transformants were selected by regeneration and rooting on medium containing kanamycin. Southern blot and PCR analysis indicated regeneranrs contain the NPTII and GUS genes. Mapping the chromosomal location of the NPTII gene is in progress.
Shoot tips of carnations (Dianthus caryophyllus L. cv. CSU White Pikes Peak) formed multiple shoots on agar nutrient medium containing 0.5 mg/liter kinetin and 0.1 mg/liter α-napthaleneacetic acid. Tissue with shoots was transferred to liquid medium on a culture wheel rotating 1 rpm. Many axillary shoots formed and eventually separated from the parent shoot. Tissue could be subcultured into fresh medium, stored at 4.5°, or rooted, potted and grown to flowering. All 175 plants flowered had normal white flowers with characteristic red flecks, indicating that the chimeral arrangement of the petal tissues had not been disturbed by the culture procedure.
Large numbers of normal chrysanthemum plants were produced from tissue cultures derived from 0.5 mm high shoot-tips. The cultures, which consisted of callus with superficial meristems, doubled in weight every 3 days in liquid Murashige-Skoog medium containing 2.0 mg/1 kinetin and 0.02 mg/1 NAA. Cultures resumed growth after at least 6 months storage at 4.5°C. Plantlets formed within 6-12 weeks when small pieces of tissue were transferred to agar media containing 0.5-2.0 mg/1 kinetin. Addition of GA3 (10 mg/1) promoted formation and elongation of leaves and shoots on the tissue. Ability to form plantlets was retained after repeated subculture; a culture started in April 1970 is still producing plantlets. One thousand plantlets were potted and grown in the greenhouse. Small (<1 cm high) plantlets initially grew more slowly than larger ones but almost all plants produced normal white flowers after 2 months short day treatment. This system could produce up to 9 × 1014 plantlets or 90 billion 15 cm plants within a year, a great increase over the number possible via conventional propagation.
Small (0.5 mm high) shoot tips of Chrysanthemum morifolium ‘Giant #4 Indianapolis White’ were grown on Murashige-Skoog medium containing various levels of kinetin, NAA, and GA3. Formation of roots, single or multiple shoots, plantlets and friable, hard or leafy callus depended on the hormone levels used. Multiple shoots and green leafy callus were produced on medium containing 2.0 mg/l kinetin and 0.02 mg/l NAA. The leafy callus was suitable for subculture and subsequent reorganization of plantlets. Multiple shoots were rooted and grown into normal plants or were used to start new cultures which formed more multiple shoots. This technique will be useful for storage and propagation of Chrysanthemum and especially for detection and rapid multiplication of virus-free plants.
Leaf pieces of pistil-sterile double petunias (Petunia hybrida Hort.) placed on a Linsmaier-Skoog (LS) medium containing 0.2 mg/liter 6-benzylamino purine (BA) produced shoots after 4 weeks which rooted within a week when the cut ends were dipped into an LS medium containing 1.0 mg/liter naphthaleneacetic acid (NAA) and then placed in a hormone-free medium. Plantlets grew rapidly after transfer to soil and had similar growth and flower type as the source clone.