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Abstract
Zygotic embryos isolated from mature berries of Vitis longii Prince and V. vinifera L. cvs. Chardonnay, French Colombard, Grenache, and White Riesling produced somatic embryos when cultured on solid Nitsch and Nitsch medium. Explants of V. vinifera cv. Pinot noir did not produce somatic embryos. The highest incidence of somatic embryogenesis occurred on medium containing (all in mg·liter−1) 1.0 NOA plus 0.2 BAP, except in ‘White Riesling’, where 1.0 NOA plus 1.0 BAP was optimal. Somatic embryos developed asynchronously from a mass of white proembryos and passed through globular, heart, and torpedo-shaped stages. Embryos proliferated by budding at the surface of globular embryos. With regular transfers, embryogenic tissues remained morphogenically competent for up to 1 year. Zygotic embryo explants cultured on medium without growth regulators usually germinated within 10 days and produced healthy seedlings. About 40% of somatic embryos germinated after isolation from their parental cultures and transfer to fresh medium. At the concentrations tested, NAA, BAP, and GA3 did not improve germination frequency over that obtained with basal medium and, in some cases, severely reduced germination. Plants were established in the greenhouse and no morphological abnormalities were observed. Chemical names used: 6-benzylam-inopurine (BAP); gibberellic acid (GA3); 2-(l-naphthyl)acetic acid (NAA); 2-naphthoxyacetic acid (NOA).
Leatherleaf fern [Rumohra adiantiformis (Forst.) Ching] fronds produced under a high-temperature regime (HTR, 30 day/25C night) grew faster and produced sori earlier than those in a low-temperature regime (LTR, 20 day/15C night). Abaxial diffusive conductance was lower for HTR-grown fronds. Light-saturated net CO2 assimilation rates (Pn) and dark respiration were lower for HTR fronds, but light-saturated Pn efficiencies (chlorophyll basis); light compensation points; and soluble sugars, starch, and nonstructural carbohydrate levels were similar for the two regimes. Transpiration and water-use efficiency (mass basis) at light saturation were similar for fronds from both temperature treatments. Comparison of physiological characteristics of fronds from the two temperature regimes revealed no differences that might account for reduced postharvest longevity of fronds produced at the higher temperatures.
Adventitious shoots developed within 3 weeks from the petiolar stub and, less often, from wounded lamina tissues when leaves excised from nodal cultures of Vitis vinifera L. cvs. French Colombard and Thompson Seedless were cultured on solid Nitsch and Nitsch medium containing BAP at 2 mg·liter-1. The youngest leaf that could be excised, from 1 to 8 mm long, was the most responsive (90% of explants producing shoots compared to 16% for leaf 6). Removal of the lamina from the petiolar stub within the first 3 weeks of culture reduced shoot production. Increase in nodal culture age, without transfer to fresh medium, had no effect on subsequent regeneration from the youngest leaves but did reduce the regeneration frequency of leaves at the next position from 43% to 20%. In regularly subculture nodal cultures, the number of transfers had no effect on subsequent regeneration. Leaves from recently established shoot tip cultures were more responsive than leaves from nodal cultures. The frequency of shoot production was higher in laterally bisected than intact leaves (70% vs. 43%) due to additional regeneration from the distal leaf half at the sites of severed veins. Shoot outgrowth was promoted by the isolation and subculture of regenerating tissue to fresh regeneration medium. Petiolar stub removal promoted de novo shoot organogenesis from the resulting lamina wound. Shoots rooted at a high frequency on Murashige and Skoog medium with 1 mg IA-A/liter and produced morphologically normal plants. Chemical names used: 6-benzylaminopurine (BAP); indole-3-acetic acid (IAA).
Abstract
Leatherleaf fern [Rumohra adiantiformis (G. Forst) Ching] fronds became desiccated rapidly after harvest with water potential decreasing from -0.45 to -1.75 MPa within 30 minutes. Partial closing of stomates 30 minutes after harvest accounted for a decrease in the rate of frond desiccation and frond water potential was -2.26 MPa after 180 minutes. Postharvest frond desiccation to water potentials of -1, -2, and -3 MPa in the field prior to water dip and cold storage resulted in frond curl of 32, 56, and 84%, respectively, when placed in postharvest rooms. Water uptake decreased during the first 4 days in postharvest rooms. Declining frond water potentials suggested reduced rate of water uptake was due to blockage of the xylem. However, no obstructions were observed at cut end of stipe from fern with frond curl or those not exhibiting frond curl. Frond water potentials were lower one hour after harvest than when undergoing normal postharvest senescence. Fronds did not exhibit normal drought-imposed wilt or frond curl during prestorage stress. Desiccation resulted in frond curl in some experiments but had little effect in others. These results indicate that frond curl can be triggered by desiccation stress but other factors are predisposing fronds to this disorder.
Broccoli and cauliflower are among the most regeneratively intractable genotypes found in the brassicaceae. To develop a method for transfer of the gene encoding S-adenosylmethionine hydrolase (SAMase) into inbred broccoli and cauliflower germplasm, we investigated the morphogenic competence and Agrobacterium susceptibility of a wide range of tissues of varied source. Appropriately controlled expression of the SAMase gene should, theoretically, reduce the plant's capacity for ethylene biosynthesis and extend the post harvest shelf life of the flower head.
Through examination of the in vitro response of a wide range of tissues we identified procedures which support caulogenesis from 100% of explants, each producing more than 30 shoots which readily convert to plantlets. Studies with several wild type and disarmed Agrobacterium strains, and utilization of the binary vector system and appropriate marker and reporter genes, led to the identification of methods for high frequency T-DNA transfer to explant tissues and the flow frequency of transgenic plants containing SAMase gene.
The developmental anatomy of direct shoot organogenesis from in vitro leaves of Vitis vinifera L. cv. French Colombard was studied by light microscopy. Regenerating petiole stubs of leaf explants were fixed at intervals and were sectioned longitudinally to determine the developmental sequence of direct shoot organogenesis. After 6 days, three distinct regions of meristematic activity were apparent within expanding petiole stub: the wound-response, organogenic, and vascularization regions. In the organogenic region, divisions of vacuolate outer cortical cells formed nodular bumps that sometimes became adventitious leaves. Promeristems, which had the potential to become adventitious shoot meristems, were also initiated asynchronously in the organogenic region. Promeristem initiation occurred by two or several synchronous cell divisions occurring in the epidermal and subepidermal cell layers. Adventitious shoots and leaves developed new vascular bundles that connected to the pre-existing vascular bundles of the explant.