Saturated and mono-unsaturated long-chain hydrocarbons from lemon juice sacs

Three varieties of lemon—Lisbon, Malta and Kusner—were examined for their content of juice sac saturated and mono-unsaturated long-chain hydrocarbons. The saturated fractions were 20 times the concentrations of the monoene fractions. The dominant linear hydrocarbon in the saturated fraction was C₂₅ while C₂₉ predominated in the monoene fraction. The saturated hydrocarbon profiles for Lisbon and Kusner were very similar to the profiles previously reported for Eureka lemon and Persian lime. The mono-unsaturated profiles were distinct for each of the three lemon varieties. In addition the lemon mono-unsaturated profiles were quite distinct from the hydrocarbon profiles previously reported for several other citrus species. In general the data support the elongation-decarboxylation mechanism for hydrocarbon synthesis proposed by Kolattukudy.     

In vitro culture of Ginkgo

Summary Ginkgo biloba L. is an important landscape tree, is resistant to insect, fungi and other pests, and produces a number of chemicals that have pharmaceutical properties (termed ginkgolides). Studies were initiated to establish an in vitro culture protocol for Ginkgo. Explants (intact embryos, embryos with cotyledons removed, and cotyledon tissue) were removed from disinfested seeds and cultured on Murashige and Skoog minimal organics medium with various combinations of either 2,4-dichlorophenoxyacetic acid (2,4-D) or naphthaleneacetic acid (NAA) and either kinetin or benzyladenine (BA). Cultures were incubated in the light and morphological development was recorded. Both embryo and cotyledon explants produced callus (cotyledon tissue produced the most callus). Ginkgolides A and B were detected in callus tissue extracts. Intact embryo cultures initiated on media with 2,4-D plus NAA for 5 wk produced shoots and roots when transferred to media with 4.5 μM 2,4-D alone for an additional 5 wk. Plants were transferred from the 2,4-D media to pots and maintained in the greenhouse.     

In vitro culture of Cedrela fissilis Vellozo (Meliaceae)

An efficient micropropagation protocol was developed for Cedrela fissilis (Meliaceae) using nodal segments from juvenile origin for axillary shoot proliferation. Shoot proliferation was significantly affected by salt formulation, explant origin and 6-benzyladenine concentration. Maximum multiplication rates (6–7 new plants were produced in the second subculture cycle per single cotyledonary node cutting) were achieved on Murashige and Skoog media supplemented with 1.25–5.0 M 6-benzyladenine. Addition of -naphthaleneacetic acid to these media caused significant inhibition on shoot proliferation and growth and stimulated callus formation. High frequency callus initiation and synergistic effects on callus growth were achieved on Murashige and Skoog medium supplemented with 6-benzyladenine at either 1.25, 2.5 or 5.0 M combined, respectively, with 2.5, 1.25–5.0 or 5.0 M -naphthaleneacetic acid. Rooting was achieved, after 10–12 days, with 87–100% of the node cuttings on half strength Murashige and Skoog medium either without growth regulators or supplemented with 2.5 M indole-3-butyric acid. Regenerated plants were successfully acclimatized on sterilized sand, for 21 days, but for further plant development the sand:soil (1:1) mixture was the best substrate. The survival rate of plantlets under ex vitro conditions was 100% after 3 months. The optimized micropropagation and callus culture protocols offer the possibility to use the organ/cell culture techniques for vegetative propagation, cryopreservation and secondary metabolism studies.     

In vitro culture of Plasmodium berghei using a new suspension system

Attempts were made to develop techniques for the continuous in vitro culture of Plasmodium berghei. The candle jar method (Trager &; Jensen, 1976) proved to be insufficient for the culture of this rodent malaria parasite. Parasitaemia decreased rapidly after the first schizogonic cycle in culture. A simple suspension technique was developed using a newly designed culture apparatus which can be placed in the laminar-flow. All manipulations necessary for the refreshment of medium and cells can be made with almost no disturbance of the culture conditions. With this system it was possible to culture P. berghei repeatedly for more than a week, completing at least four schizogonic cycles with considerable mcrozoite invasion and a 2–6-fold multiplication. Infection rates of up to 6.0% were achieved and cultures were maintained for 9 days. Several specific properties of P. berghei and the differences between the candle jar method and the new suspension method are discussed to explain the results obtained in both systems.     

In vitro culture of Phyllanthus caroliniensis (Euphorbiaceae)

An efficient micropropagation protocol was developed for the medicinal plant Phyllanthus caroliniensis (Euphorbiaceae) using nodal segments for axillary shoot proliferation. Maximum multiplication (21–23 shoots per explant) was achieved on MS or AR media supplemented with either 5.0 μM BA, 1.25–5.0 μM kinetin or 2.5–5.0 μM 2iP. Rooting was achieved with 80–100% of the microshoots on MS medium without growth regulators, although 1.25 μM NAA and 1.25–5.0 μM IAA promoted significant increases in the number of roots per explant. Regenerated plants were successfully acclimatized and about 88% of plantlets survived under ex vitro conditions. Flowering was observed on in vitro grown plantlets and after 3–4 weeks of acclimatization. High frequency callus initiation and growth was achieved when nodal segment explants were inoculated in the vertical position on MS medium supplemented with 5.0 μM 2,4-D. Root cultures were successfully established on MS medium containing 1.1 μM NAA. The optimized micropropagation, callus and root culture protocols offer the possibility to use cell/root culture techniques for vegetative propagation and secondary metabolism studies. This revised version was published online in June 2006 with corrections to the Cover Date.     

In vitro culture and plant regeneration of large flowered purslane

Culture conditions were established for callus induction from a range of Portulaca grandiflora Hook tissues. Rapidly growing calli were obtained on Murashige and Skoog medium with stem-, leaf- and sepal-derived explants. Plant regeneration via organogenesis was explant-origin dependent with hypocotyl tissues giving the highest shooting frequency. Light conditions, pH and carbon source had a pronounced effect on the percentage of explants regenerating buds and the number of buds formed. It was possible to establish stable regenerated plants in the glasshouse.Abbreviations BA 6-benzyladenine - 2,4-D 2,4-dichlorophenoxyacetic acid - NAA 1-naphthaleneacetic acid - IAA indoleacetic acid - MS Murashige and Skoog (1962) medium     

In vitro meristem culture of M.4 apple (Malus pumila Mill.). I. Optimal nutrient medium

Shoot tips of M.4 apple clone were excised from actively growing one year-old stoolbed branches, and cultured in order to determine the optimal nutrient medium for each stage of their in vitro culture. The basal medium (BM) used was that described by Murashige and Skoog, supplemented with vitamins, glycine, myoinositol, sucrose, with or without agar, and different combinations of plant growth regulators. Best media for each stage were: BM+0.5 mg 1^-1 indole-3yl-butyric acid (IBA)+0.5 mg 1^-1 6-benzylaminopurine (BAP) for explant establishment (Stage I); BM+0.1 mg 1^-1 IBA+1.0 mg 1^-1 BAP for multiplication and internode enlargement (Stage II); and 2.0 mg 1^-1 IBA+0.1 mg 1^-1 BAP without agar for the rooting of the plantlets (Stage III).     

In vitro culture of Safflower L. cv. Bhima: initiation,growth optimization and organogenesis

Callus induction and in vitro plantlet regeneration systems for safflower (Carthamus tinctorius L.) cv. Bhima using root, hypocotyl, cotyledon and leaf explants were optimized by studying the influence on organogenesis of seedling age, media factors, growth regulators and excision orientation. Supplementation of the medium with an auxin: cytokinin ratio < 1 enhanced the growth rate of callus cultures; however, for 2,4-D the ratio was > 1.34–11.41 μM concentrations of growth regulators (IAA, NAA, BA and Kinetin) in the medium were found effective for callus induction and regeneration in all explants. The calli could be maintained over 32 months. BA (4.43 μM) combined with casein hydrolysate (10 mg l-1) yielded the highest rate of shoot production on hypocotyl (3–6) and cotyledon (5–7) explants and cotyledonary derived callus (4–8). More shoots were produced on explants cut from the most basal region of cotyledons from 5 to 7-day-old seedlings than from older seedlings or more distal cut sites. Apolar placement of explants, inhibited shoot regeneration. The shoot regeneration potential remained upto 7 months in calli developed on NAA + BA. Of three media tested, MS was superior to SH-M and B5. Rooting of shoots was not efficient; 42% of the shoots were rooted on MS medium containing sucrose (7–8%) + IAA (2.8–5.7 μM). Capitula induction was observed in both callus mediated shoots on cotyledons and shoots on rooting medium with sucrose, IAA, NAA and IBA. Well developed plantlets were transferred to the field with a 34% success rate. This revised version was published online in June 2006 with corrections to the Cover Date.     

白刺组织培养技术的研究

试验选用唐古特白刺幼嫩茎段和叶片作为材料,研究白刺不同外植体的离体培养技术。结果表明,白刺带芽嫩茎是诱导丛生芽的良好外植体,而叶片是诱导愈伤组织的良好外植体;白刺的最适增殖、壮芽培养基是：MS BA0.5mg／L NAA1．0mg／L GA32．0mg／L;最适生根培养基是：l／2MS IBA0.5mg／L;愈伤组织诱导培养基是：MS 2,4-D0.5～1．0mg／L。     

Pectins from the albedo of immature lemon fruitlets have high water binding capacity

The white part of citrus peel, the albedo, has a special role in water relations of both fruit and leaves from early on in fruit development. In times of drought, this tissue acts as a water reservoir for juice sacs, seeds and leaves. When water was injected into the albedo, free water was undetectable using magnetic resonance imaging. Microscopy showed tightly packed cells with little intercellular space, and thick cell walls. Cell wall material comprised 21% of the fresh albedo weight, and contained 26.1% galacturonic acid, the main constituent of pectin. From this, we postulated that pectin of the cell wall was responsible for the high water-binding capacity of the immature lemon albedo. Cell wall material was extracted using mild procedures that keep polymers intact, and four pectic fractions were recovered. Of these fractions, the SDS and chelator-soluble fractions showed viscosities ten and twenty times higher than laboratory-grade citrus pectin or the other albedo-derived pectins. The yield of these two pectins represented 28% of the cell walls and 62% of the galacturonic acid content of immature lemon albedo. We concluded that, from viscosity and abundance, these types of pectin account for the high water-binding capacity of this tissue. Compositional analyses showed that the two highly viscous pectic fractions differ in galacturonic acid content, degree of branching and length of side chains from the less viscous albedo-derived pectins. The most striking feature of these highly viscous pectins, however, was their high molecular weight distribution compared to the other pectic fractions.     

In vitro culture of shoots of Pinus caribaea on a liquid medium

Shoot apices of a clone of Pinus caribaea Morelet were cultured and multiplied in vitro by supporting them with their basal cut ends immersed in a liquid nutrient medium.The initial heights of explants and their initial numbers of leaves were positively correlated with the numbers of buds and shoots produced by the explants after a bud induction phase and after a shoot elongation phase. The final numbers of buds and shoots were positively correlated with reductions in the quantities of phosphorus detected in the media and negatively correlated with the numbers of brown leaves produced on the explants.In a comparison between the growth of shoot explants on liquid and solid media, shoots incubated on the liquid medium showed significantly greater increases in length in a four-week period than those cultured on solid medium.This technique, using liquid media, provides a system in which both the nutrient utilization and the growth rates of isolated pine tissues can be readily assessed. Furthermore, the multiplication rate of the tissue can be predicted following the observation of correlated characters early in the micropropagation cycle.     

In vitro development of one-cell embryos from outbred mice: Influence of culture medium composition

Summary One-cell embryos from outbred mice (CF1, CD-1, and Dub:ICR) were cultured in various modifications of egg culture medium (ECM). The best development was observed in medium in which inorganic salts of modified T6 medium (mT6) replaced those of ECM. In this modification (TE), 66% of one-cell CF1 embryos developed into blastocysts, comared to 46 and 43% for ECM and mT6, respectively. Moreover, the cell numbers of blastocysts developing in TE (74.9±3.3) were higher than the cell numbers of those developing in ECM (55.1±2.4). The culture requirements of embryos varied between different stocks of mice: Fewer CF1 embryos developed to the blastocyst stage than either Dub:ICR embryos (90%) or CD-1 embryos (84%). Lowering the osmolarity of the medium from 300 to 280 mOsm, increasing the concentration of KC1 from 1.42 to 25 mM, or omitting lactate from the medium during Day 1 of culture did not further improve development of embryos, in contrast to previous reports. However, the time at which embryos were transferred to outgrowth medium influenced their postblastocyst development. The best development was observed when embryos were transferred on Day 4 of culture at the late morula-early blastocyst stage. This work was supported by the Office of Health and Environmental Research, U.S. Department of Energy, Washington, DC, contract DE-AC03-76-SF01012.     

In vitro growth of embryos and callus of coconut palm

Summary A medium for optimal growth of embryos of Jamaican Tall and Green Malayan Dwarf varieties of coconut palm was developed. The liquid basal Murashige and Skoog medium was supplemented with coconut milk, IAA and 2IP. Activated charcoal improved embryo growth on agar medium. A single callus line was initiated from solid endosperm and subcultured on basal Schenk and Hildebrandt medium supplemented with 2 mg per 1 NAA. Attempts at inducing organogenesis in the callus were unsuccessful. No vascular tissue was present. The callus was aneuploid with the chromosome number=8 (normal 2n=32). Florida Agricultural Experiment Stations Journal Series No. 542. The research was supported in part by the Horticultural Research Institute (to J. H. T.) and the American Philosophical Society (to J.B.F.).     

The effect of different irrigation treatments on yield and quality of Verna lemon

Verna lemon trees were irrigated by six different treatments: five flood and one drip. Soil and plant water status, yields and fruit quality were measured. The drip-irrigated treatment gave higher yield and fruit size. The levels of soluble solids, acidity and sugars in the lemon juice decreased in the treatments that used most water. This is explained by a dilution effect. An erratum to this article is available at .     

驱蚊草组织培养及其愈伤组织诱导研究

运用正交设计方法,利用植物组织、细胞培养技术,成功地建立了驱蚊草组织培养快繁技术体系。研究出:不定芽诱导的最适培养基为MS 6-BA0.2mg/L NAA0.3mg/L,增殖倍数为6.73;生根培养基为1/2MS NAA0.4mg/L,平均每株生根数为11.2条,生根率为90.0%。通过对其离体茎段的培养研究实验,得出驱蚊香草的最适愈伤组织诱导培养基为MS 2,4-D0.5mg/L 6-BA2.0mg/L NAA0.3mg/L,发愈率为93.3%,愈伤组织多为淡黄色,质地疏松。     

In vitro culture of Cucumis sativus L. XVIII. Plants from protoplasts through direct somatic embryogenesis

A procedure is described for the isolation and culture of protoplasts from embryogenic callus (gel-like callus — GLC) and embryogenic suspension cultures (ESC) of Cucumis sativus c.v. Borszczagowski. Maximal protoplast yields from GLC and ESC were 5×10⁶ and 1×10⁷ protoplasts/g tissue respectively. They were obtained following 14–16 h digestion with 1.2% Cellulase Onozuka R-10, 1.2% Macerozyme R-10 and 0.3% Driselase. At a plating density of 2×10⁵ / ml, first divisions occurred in 4–5 days and 7–8 days in ESC-and GLC-derived protoplasts respectively. The highest percentage of direct embryogenesis (over 80%) was observed with ESC. It was possible to obtain approximately 5000 embryo structures / g tissue. Some embryos converted into plants after 6 weeks, but most of them after 2 months of culture. ESC-derived plants, when transferred into the glasshouse, bloomed normally, and set seeds.Abbreviations CMS Murashige & Skoog (1962) medium for cucumber - GLC gel-like callus - ESC established embryogenic suspension culture - 2,4-d 2,4-dichlorophenoxyacetic acid     

In vitro interaction of zajdela ascites hepatoma cells with lipid vesicles

We studied the in vitro interaction between Zajdela ascites hepatoma cells and small unilamellar vesicles, consisting of ¹⁴C-labeled phosphatidylcholine, cholesterol, and phosphatidylserine (molar ratio 5: 4: 1), containing high intravesicular concentrations of carboxyfluorescein or fluorescein isothiocyanate tagged dextran.The entrapped markers were found to be associated with the cells to a lesser degree than the vesicle membrane marker. This discrepancy, which is slightly less pronounced for fluorescein isothiocyanate tagged dextran than for carboxyfluorescein, increases with incubation time and decreases with increasing vesicle lipid concentration in the incubation mixture. Vesicle-plasma membrane exchange of the vesicle lipid marker could not entirely explain the observed discrepancy. It is tentatively concluded that the gap mainly arises from a selective loss of entrapped dyes from vesicles actually interacting with the cell surface. Both spectrofluorimetric and fluorescence microscopic observations, as well as the relative insensitivity of vesicle uptake towards the presence of metabolic inhibitors, exclude a major contribution of endocytosis as a vesicle uptake route. We therefore conclude that vesicles are primarily internalized by a vesicle-cell fusion-like process. The observed discrepancy in uptake between entrapped materials and vesicle lipid is discussed in terms of a two-site vesicle cell surface interaction model.     

In vitro propagation of Amaryllis belladonna

Amaryllis belladonna L. plants were multiplied successfully by means of tissue culture techniques. Different plant parts were tested as explant material, but plantlets could only be generated from the twin-scales and immature scapes. These in vitro-formed plantlets were divided into four parts and used for further multiplication. The twin-scale explants had the highest multiplication rate when a medium with 22.2 M benzyladenine and 0.54 M naphthaleneacetic acid was used. The sucrose concentration played an important role in the initiation of new plantlets, and the best results were obtained when a sucrose concentration of 2–3% was used. Anatomical observations were made during the initiation of the new plantlets.Abbreviations BA benzyladenine - NAA naphthaleneacetic acid - Benomyl (methyl [1-[(butylamino) carbonyl]-1H-benzimidazol-2-yl] carbamate) - Folpet (2-[(trichloromethyl)thio]-H-isoindole-1,3(2H)-dione phthalimide(I))     

In vitro regeneration of long spur barrenwort (Epimedium grandiflorum Morr.) from rachis explants

Summary A system for micropropagation of Epimedium grandiflorum Morr. from rachis explants was developed. Explants were cultured onto Murashige and Skoog (MS) basal salts medium supplemented with (per L) 100 mg myo-inositol, 2 mg pyridoxine-HCl, 2 mg nicotinic acid, 0.40 mg thiamine-HCl, 30 g sucrose, and 2 g Phytagel. The medium also contained 2,4-dichlorophenoxyacetic acid (2,4-D) at 0.1, 0.2, or 0.25 mg/L (0.5, 0.9, or 1.1 μM) combined with either N⁶-benzyladenine (BA) or 2-isopentenyl adenine (2ip) at 2.5, 5, or 10 mg/L (11.1, 22.2, or 44.4 μM BA or 12.3, 24.6, or 49.2 μM 2iP). Cultures were maintained at a 16-h photoperiod (40 μmol/m²/s) and 23±2° C. Callogenesis preceded shoot regeneration. Callus formation increased with higher 2,4-D concentrations. The highest percent regeneration, 83% of explants, was obtained on 10 mg BA per L (44.4 μM) combined with 0.25 mg 2,4-D per L (1.1 μM). The maximum number of shoots, 15 per explant, was obtained from explants cultured on a medium containing 0.1 mg 2,4-D per L (0.45 μM) combined with 2.5 mg BA per L (11.1 μM). Maximum shoot length, 0.4 cm, was obtained on 5 mg BA per L (22.2 μM) combined with 0.2 mg 2,4-D per L (0.9 μM). To produce whole plants, shoots were separated and rooted on hormone-free medium containing 1 g activated charcoal per L. Rachises provided an excellent source of explants for Epimedium micropropagation and proved suitable for callus production.