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  • Author or Editor: A. Raymond Miller x
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A protocol was developed for excising and culturing cotyledon explants from mature achenes of strawberry (Fragaria × ananassa Duch.). Cotyledon explants formed callus with multiple shoot buds on agar-solidified Murashige and Skoog media containing several combinations of hormones (1 μm 2,4-D; 10 μm 2,4-D; 1 μm BA + 1 μm 2,4-D; 1 μm BA + 10 μm 2,4-D; 5 μm BA; 5 μm BA + 1 μm 2,4-D; 5 μm BA + 10 μ m 2,4-D; 5 μ m BA + 5 μm NAA; 5 μ m BA + 15 μ m NAA). After three subcultures, only tissues maintained on the medium containing 5 μm BA + 5 μm NAA continued to form shoots. Tissues transferred to other media eventually died (1 μm 2,4-D; 1 μ m BA + 10 μ m 2,4-D; 5 μ m BA; 5 μ m BA + 1 μ m 2,4-D), became unorganized (1 μm BA + 1 μm 2,4-D; 5 μm BA + 10 μm 2,4-D; 5 μm BA + 15 μm NAA), or formed roots (10 μm 2,4-D). Whole plantlets were produced by transferring callus with buds to medium lacking hormones. The rapid regeneration of clonal plantlets from cotyledon explants may be useful for reducing variability in future developmental studies. Chemical names used: N-(phenylmethyl)-1H-purin-6-amine (BA); (2,4-dichlorophenoxy) acetic acid (2,4-D); and 1-naphthaleneacetic acid (NAA).

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A factorial combination of pretreatments, fixatives, and stains was examined to identify the best available method for staining the chromosomes of strawberry (Fragaria × ananassa Duch. `Chandler') root-tip cells. Three pretreatments (a -bromonaphthalene, p -dichlorobenzene, and 8-hydroxyquinoline), three fixatives (Farmer's fluid, Carnoy's fluid, and Newcomer's fluid), and five stains (acetocarmine, alcoholic hydrochloric-acid carmine, altered carbol fuchsin, lacto-propionic orcein, and leucobasic fuchsin) were tested. Pretreatment with either a -bromonaphthalene (saturated aqueous) or 8-hydroxyquinoline (2 mm) for 5 hours at 14C, overnight fixation in Farmer's fluid, hydrolysis in 1 n HCl (15 minutes at 60C), and staining with altered carbol fuchsin produced chromosome preparations superior to other treatment combinations. Treatment with a-bromonaphthalene, Farmer's fluid, and alcoholic hydrochloric-acid carmine (2 days at 25C) also produced acceptable chromosome preparations.

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A comparison of pretreatment, fixing, and staining methods for root tips of Fragaria × ananassa (2n=8x=56), a polyploid species with small chromosomes, was made to facilitate chromosome counting. Three pretreatments (8-hydroxyquinoline, α-bromonaphthalene, and p-dichlorobenzene), three fixatives (Farmer's, Carnoy's, and Newcomer's), and five stains (acetocarmine, lacto-propionic orcein, leucobasic fuchsin, altered carbol fuchsin, and alcoholic hydrochloric-acid carmine) were examined in a factorial design to determine which treatment combination produced the best chromosome preparation. Field propagated runners were grown in sand under greenhouse conditions with supplemental lighting to produce root tips for late morning collection. The treatment combinations of α-bromonaphthalene or 8-hydroxyquinoline, Farmer's fixative, and altered carbol fuchsin, or the combination of α-bromonaphthalene, Farmer's fixative, and alcoholic hydrochloric-acid carmine produced the most intensely-stained and well-defined preparations.

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F1 hybrids between high leptine-producing clones (8380-1, PI 458310 and 55-1) of Solanum chacoense Bitt. and anther culture competent or anther-derived clones of S. phureja Juz. & Buk. that did not produce leptines were generally weak plants that grew slowly and died before flowering. Exceptional hybrids could be found that were capable of completing a life cycle, especially during the hot summer months in the greenhouse. All F1 hybrids produced leptines in the leaves but not the tubers, albeit at lower levels than in the S. chacoense parent. Anther-derived monoploids from the F1 hybrids exhibited a range of leptine production from none to levels approaching the S. chacoense parent. Backcross populations of an F1 hybrid to the S. chacoense and S. phureja parents were examined for leptine production. Backcross hybrids were generally much more vigorous than the F1 hybrids. All of the S. chacoense backcrosses produced leptines ranging from intermediate to high levels; four of the twelve S. phureja backcrosses exhibited low leptine levels. A general dominance of leptine synthesis was therefore exhibited, although the nonleptine-producing parent affected the expression of leptines in the hybrids.

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A sequential study was undertaken to optimize the cultural conditions for potential tetrahaploid plant regeneration from anthers of `Chandler', `Honeoye', and `Redchief' strawberries. A comparison of auxins (IAA/NAA), cytokinins (BA/BPA/KIN) and carbohydrates (suc/glc/mal) showed highest regeneration with 2mg/l IAA, 1mg/l BA and 0.2M glc in agar-solidified MS medium after 60d (30d darkness + 30d 16h white light photoperiod). When comparing MS. Nitsch & Nitsch, and H1 (a new formulation based on the anther culture literature) media containing IAA/BA/glc solidified with Phytagar® or Gelrite®, highest regeneration occurred with H1 with Gelrite. Lastly, Fe-EDTA in H1 medium with IAA/BA/glc yielded more shoots than H1 medium containing Fe-Metalosate®, and anthers cultured in darkness for 30d followed by 30d in white light produced more shoots then those cultured in white or yellow light (16h photoperiod) for the initial 30d. Hence, the optimum regeneration medium and conditions for shoot regeneration from anthers of these 3 cultivars was H1 containing 2mg/l IAA, 1mg/l BA, 0.2M glc, Fe-EDTA, and Gelrite, followed by incubation in darkness for the first 30d. Although differences were noted for regeneration efficiency between cultivars, initial chromosome counts confirmed that tetrahaploids (n=4x=28) were obtained.

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Abstract

Peroxidase activity was determined in cucumber fruit (37 to 55 mm in diameter) subjected to mechanical stress followed by storage at 25C and 100% RH. Compared to unstressed, unstored control fruit, severe mechanical stress stimulated total extractable peroxidase activity after 24 and 48 hr of storage. Moderate mechanical stress or storage alone stimulated peroxidase activity to lesser degrees. Peroxidase isozyme analysis by native polyacrylamide gel electrophoresis showed that cucumber anodic peroxidases could be separated into slow-, moderate-, and fast-migrating groups. Both moderate and severe mechanical stress caused the appearance of a new slow-migrating peroxidase isozyme immediately after treatment. This slow-migrating isozyme disappeared after 24 hr of storage, then reappeared after 48 hr. Severe stress induced the appearance of two additional moderate-migrating peroxidases 24 hr after treatment. The same fast-migrating isozymes were also present in unstressed and moderately stressed fruits stored for 48 hr. Hence, using peroxidase as an indicator, it appears that mechanical stress induces an accelerated aging of processing cucumber fruit.

Open Access

Peroxidase activity in extracts from freeze-dried tissue of Fragaria × ananassa Duch. cv. Chandler was highest in tissue-cultured (TC) plants, followed by field-grown (FG) and lowest in greenhouse (GH) plants. Among tissue types, activity was highest in petioles, with leaves second highest. Fruit, root, and crown tissue all exhibited low or no activity. When subjected to isoelectric focusing (IEF), petiole tissue extracts exhibited more isozymes than extracts from other organs regardless of staining substrate. Using 4-chloro-1-naphthol and H2O2 as substrates, anionic and cationic isozymes were observed in TC petiole extract with nine isozyme bands ranging in pI from 3.9 to 9.5. In TC leaf extract an isozyme at pI 7.4 was observed that was not present in other organ extracts when H2O2 and benzidine, p-phenylenediamine or 3-amino-9-ethylcarbazole were used as substrates. Specific isozymes and number of isozymes varied according to plant organ and developmental stage. Mature leaves and over-ripe fruit appeared to exhibit more activity and a larger number of isozymes than developing tissues of those plant organs.

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Mechanical stress received by pickling cucumbers (Cucumis sativus L.) during harvest can cause physiological degeneration of the placental tissues, rendering the cucumbers unsuitable for use in some pickled products. Cucumbers were subjected to controlled stresses by dropping and rolling under weights to induce such degeneration. Following storage at various temperatures for O, 24, and 48 hours, refreshed delayed light emission from chlorophyll (RDLE) was measured and transmission electron micrographs of chloroplasts were made. Mechanical stress rapidly suppressed RDLE and induced accumulation of starch granules within the chloroplasts. Rolling usually had a greater effect on RDLE than did dropping. After 48 hours, RDLE suppression persisted; starch granules were no longer evident in chloroplasts from mechanically stressed fruit, but very electron-dense inclusions had developed in the chloroplasts. Storage temperatures affected RDLE levels but had minimal interaction with stress responses. Cucumber lots subjected to excessive mechanical stress likely could be detected using RDLE measurement.

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A nondestructive method was developed utilizing a modified Trebor 101 watercore tester to evaluate the internal quality of pickling cucumbers. The method involved measuring the relative amount of visible-infrared light passing through the longitudinal midsection of whole cucumber fruit. Light transmission was quantified on a unitless sigmoid scale from 1 to 10, with light transmission and scale values positively related. Immediately after hand harvest, size 3F (47 to 51 mm in diameter) cucumbers exhibited transmission values between 2 and 3, regardless of cultivar. Following a mechanical-stress treatment, which simulated bruising incurred during harvesting and handling of cucumbers, the internal quality of the fruit declined and was associated with an increase to a value of 6 in light transmission compared to non-stressed fruit. Light transmission increased as the severity of stress applied to the fruit increased, and high light transmission values were evident throughout a 48 h storage period at room temperature. Light transmission values increased as fruit diameter decreased, but values within a particular size class of undamaged, hand-harvested fruit were consistent. Machine-harvested fruit (size 3F), evaluated just before processing, exhibited light transmission values from 2 to 8, but the majority of fruit fell within the transmission range of 2 to 3. When fruit exhibiting different light transmission values were speared (cut longitudinally into sixths), processed, and then visually evaluated by panelists, spears prepared from fruit exhibiting high transmission values were judged to be of lower quality than those prepared from fruit exhibiting low transmission values. Visible-infrared light transmission may be a valuable tool for detecting poor quality cucumbers before processing, and could allow the mechanical selection of high quality fruit on a large scale basis.

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Abstract

Cucumber (Cucumis sativus L.) fruit (‘Heinz 3534’) subjected to mechanical stress followed by storage for 48 hr exhibited visible degeneration of the mesocarp and endocarp, which was accompanied by several-fold increases in the activity of the enzymes pectin methylesterase, peroxidase, polygalacturonase, and xylanase. The activity of all these enzymes increased in the endocarp, whereas only pectin methylesterase and polygalacturonase increased in the mesocarp, and pectin methylesterase, peroxidase, and polygalacturonase increased in the exocarp. Further, the increase in the activity of pectin methylesterase, peroxidase, and polygalacturonase was less when cucumbers were stored at 0° or 10°C vs. 25° or 38° after mechanical stress. Cucumbers stored for only 8 hr after mechanical stress, or not stressed, and stored for 8 or 48 hr showed no consistent significant increases in enzyme activity. Endocarp firmness of fruit stored at 25° or 38° for 8 hr after mechanical stress was lower than that of unstressed fruit, but this decrease was not evident after 48 hr of storage, and mesocarp firmness was not affected by mechanical stress regardless of storage temperature or time. Ethylene production was stimulated significantly by 8-hr storage at 0°, following mechanical stress, and by 48-hr storage at 0°, but was unaffected by all other treatment and storage regimes. These data indicate that mechanical stress induces biochemical and morphological changes in the major tissues of cucumber fruit, but tissue firmness and/or ethylene production will not serve as indicators of these changes. Moreover, the effects of mechanical stress do not appear to be mediated through the action of ethylene.

Open Access