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  • Author or Editor: Mark P. Bridgen x
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Traditional and biotechnological breeding techniques are being united to develop exciting new plants and to improve existing cultivated plants by introducing natural variability from germplasm resources. Intervarietal, interspecific and intergeneric crosses can be accomplished by using plant embryo culture techniques, sometimes also referred to as embryo rescue. Embryo culture involves the isolation and growth of immature or mature zygotic embryos under sterile conditions on an aseptic nutrient medium with the goal of obtaining a viable plant. The technique depends on isolating the embryo without injury, formulating a suitable nutrient medium, and inducing continued embryogenic growth and seedling formation. The culture of immature embryos is used to rescue embryos from hybrid crosses that were once thought to be incompatible because they would normally abort or not undergo the progressive sequence of ontogeny. The culture of mature embryos from ripened seeds is used to eliminate seed germination inhibitors, to overcome dormancy restrictions, or to shorten the breeding cycle. New and exciting cultivars of Alstroemeria, also known as Lily-of-the-Incas, Inca Lily, or Peruvian Lily, have been bred by using zygotic embryo culture; these techniques and applications will be discussed.

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In vitro growth and development of Alstroemeria `Cornell Pink' were evaluated on media containing different amounts of CaCl2, MgS O4, FeSO4, NO3, or NH4. Six levels of calcium chloride were originally examined (from 0 to 75 mM); the low levels proved to be most beneficial. Subsequent experiments used CaCl2 levels from 0 to 3.0 mM. Again, the low levels were most productive. Two experiments, with different gelling agents, were designed for MgSO4. The levels ranged from 0 to 15 mM. The 15 mM level produced explants with the greatest fresh weight. Three experiments were used to study the effect of FeSO4. The range was the same in all of the experiments (0 to 1 mM), but the increments and the gelling agents differed. In all three experiments, the 1 mM level proved to be toxic. The group with treatments from 0.01 to 0.5 mM had the best response over time. Both experiments with nitrogen found no response to different NO3:NH4 ratios. A positive linear response to rate was found within the range studied (20 to 80 mM).

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Red- and pink-flowering cultivars of Alstroemeria were grown with and without cooling tubes placed in the growing medium. Cooling tubes were placed immediately below, 5 cm below, and 10 cm below the surface of the medium. Cooling was obtained by circulating well water (10 to 15C) through polyethylene tubing. Air and media temperatures were recorded every 3 to 10 days in the morning and in the late afternoon to determine if positioning of the cooling tubes had an effect on the medium temperature. The air temperature was warmer than the noncooled medium, which, in turn, was significantly warmer than the cooled medium. There were no significant differences in media temperatures among the three cooling treatments. Flowering of cooled Alstroemeria cultivars continued for 2 months longer at the end of the summer and reinitiated 1 month earlier than the noncooled control. Both cultivars produced significantly more total flowers when grown in cooled medium. Flower production was greatest in the summer for plants with tubes 10 cm below the surface and least for plants in noncooled medium. This trend continued when flowering was reinitiated; however, by April of the following year, all treatments were equal in flower production.

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Presowing treatments and temperature regimes were tested to improve germination of Alstroemeria hybrids 3 to 12 months following harvest. In addition, seeds from 20 intraspecific F1 hybrids of five selections were also tested 3 to 7 or 8 to 12 weeks following harvest. Seeds were pretreated by chipping the seedcoat above the embryo, general abrasion of the entire seedcoat, or soaking 12 hours in distilled water, GA, (0.029, 0.29, 2.9 mm), or KNO3 (0.5 and 1.0 m). Pretreatments were evaluated under three environmental regimes: 8 weeks at a constant 18-25C (warm), 4 weeks at 18-25C followed by 4 weeks at 7C (warm-cold), or 4 weeks at 7C followed by 4 weeks at 18-25C (cold-warm). There was an interaction between pretreatment and environmental regime for percent germination. Germination percentages for the water soak and GA, at 0.29 or 2.9 mm were significantly higher than for the other pretreatments, but were not significantly different from one another. The warm-cold environment yielded higher germination percentages than the other environments. The time to germination was longest for the cold-warm regime. This response depended on the genotype and the age of the seed. Chemical name used: gibberellic acid (GA3).

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Meristems from three different positions were excised from in vitro plants of Alstroemeria genotype A30. Explants were removed from the most-distal vegetative shoot apical meristems, rhizome tip apical meristems, and rhizome tip axillary meristems. Meristems were cultured on four different media to compare the effect of meristem position and medium on the ability to produce Alstroemeria rhizomes from meristems. The meristem culture media were Murashige & Skoog salts plus 8.39 μM pantothenic acid, 1.19 μM thiamine, and 0.55 mm myo-inositol (MSM), MSM plus 8.88 μM of 6-benzylaminopurine (BA), MSM plus 8.88 μM BA, and 0.72 μM gibberellic acid (GA3), and MSM plus 0.72 μM GA3. Meristems that were removed from the vegetative shoot apices did not develop rhizomes on any medium. Rhizome tip apical meristems developed less than 10% rhizomes when subcultured on media containing BA and GA3. However, rhizome tip axillary meristems developed rhizomes on all media with best results achieved when the medium was supplemented with BA.

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Alstroemeria, the Inca Lily or Lily-of-the-Incas, is becoming a popular garden plant in the United States. In past years, the primary interest in Alstroemeria has been for its cut flowers. However, recent cold-hardy introductions (USDA hardiness zone 5) have expanded the interest of this colorful plant as a garden perennial throughout the United States. Previously, garden interests were restricted to warmer zones in the southern United States where Alstroemeria could overwinter. This research describes a breeding procedure that has been used with the objective to develop a cold-hardy, white-flowered Alstroemeria. The interspecific hybrids were bred with the use of in ovulo embryo rescue. Reciprocal crosses were made between several white-flowered cultivars and the cold-hardy Chilean species Alstroemeriaaurea during Summer 2004. Ovaries were collected 10–23 days after hand pollination and their ovules were aseptically excised. Ovules were placed in vitro on 25% Murashige and Skoog (MS) medium under dark conditions until germination. Three weeks after germination, they were then placed on 100% MS medium, and subcultured every 3–4 weeks thereafter until they were large enough for rooting. After rooting and acclimation, plants were transferred to the greenhouse. Successful hybrids that were produced in 2004 were evaluated under greenhouse and field trials during 2005, and the number of plants with white-colored flowers was noted. Although certain morphological characteristics indicate if plants are coldhardy, the hybrids will be overwintered outside in Ithaca, N.Y. (USDA zone 5), during the next several years to determine winter hardiness.

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Disinfected midrib sections of Mussaenda `Queen Sirikit' ≈3 to 4 mm in size were cultured on a basal medium of Murashige and Skoog salts and vitamins, 87.7 mm sucrose, and 5 g Sigma agar/liter supplemented with several concentrations of indole-3-acetic acid (IAA) (0, 5.0, 10.0, 20.0 μm) and 6-benzylaminopurine (BAP) (0, 0.5, 1.0, 2.5, 5.0, 10.0, 25.0, 50.0 μm). Cultures were subculture onto the same treatment after 5 weeks and observed weekly for 15 weeks for the presence of somatic embryos. As somatic embryos were produced, they were subculture onto basal medium supplemented with 0.5, 1.0, 2.5, or 25.0 μm BAP. Callus was first observed at 2 weeks in cultures grown on basal medium supplemented with 5.0–20.0 μm IAA and 0–50.0 μm BAP. Somatic embryos were observed at 8 weeks on basal medium supplemented with 5.0–10.0 μm IAA and 2.5–5.0 μm BAP. Callus cultured on 0–10 μm IAA and 5.0–10.0 μm BAP produced the greatest number of somatic embryos by 15 weeks. Somatic embryos subculture to basal medium supplemented with 25.0 μm BAP proliferated shoots, while eliminating BAP from the medium resulted in root and callus production. Shoots and entire plants were removed from in vitro conditions and successful] y acclimated to greenhouse conditions. Somatic embryo-derived plants flowered sporadically 25 to 35 weeks after removal from in vitro conditions. Variations in sepal number and leaf number per node were observed at 1% to 5%.

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