Plant regeneration from cell suspension cultures of Vigna aconitifolia

Plant regeneration has been achieved routinely from established cell suspension culture lines of Vigna aconitifolia (moth bean), a highly drought tolerant grain legume. The cultures originated from three-week-old leaf callus. Several media including MS, B₅, AA, SL, PCM, SH and L-6 were tested for their effects on cell growth. Maximum growth was observed in L-6 medium containing 44.5 M 2,4-D. After 6 to 8 weeks the suspensions were filtered through 500, 250, 125 and 60 m sieves, respectively, for four to five subcultures. An embryogenic cell line (VA-686) was obtained from the cell fraction collected below 250 m. The VA-686 cell line is being maintained on L-6 medium with 4.5 M 2,4-D and 2.3 M Zeatin. Somatic embryogenesis was induced by transferring the cells to L-6 medium with 4.6 M zeatin in which green cell clusters were produced. The somatic embryos developed from most of the cell clusters when plated on L-6 agar medium with 2.3 M BA.Plantlets were obtained from the embryos on L-6 medium with 10.0 M IBA. The regenerated plants were grown to maturity in the greenhouse.Abbreviations BA Benzyladenine - 2,4-D 2,4- dichlorophenoxyacetic acid - GA₃ Gibberellic acid - IBA Indole-3-butyric acid - IPA Isopentenyladenine - KN Kinetin - NAA Napthaleneacetic acid - AA Toriyama and Hinata, 1985 - SL Phillips and Collin, 1980 A project sponsored by United States Agency for International Development, Washington D.C.     

Nuclear pore distribution and relation to adjacent cytoplasmic organelles in cotyledon cells of developing Vicia faba

Following a zinc iodine-osmium tetroxide fixation, nuclear pore distribution was studied in 0.3-m sections from cotyledons of developing Vicia faba L. Localised absence of nuclear pores was found to be associated with proximity of organelles to the nucleus. Golgi cisternae and mitochondria are associated with areas of pore absence while cisternal endoplasmic reticulum and tubular endoplasmic reticulum are linked with areas showing reduction in pore density. Pores were seen in the nuclear membrane adjacent to vacuoles. Pattern analysis of pore distribution indicated possible clustering within an overall regularity.Abbreviations ER endoplasmic reticulum - ZIO zinc iodine-osmium tetroxide     

Direct somatic embryogenesis through thin cell layer culture in Panax ginseng

Direct somatic embryos were differentiated on cotyledon transverse Thin Cell Layers (tTCLs) of Panax ginseng after 9 weeks in the Murashige and Skoog basal (MS) medium containing 2,4-d (5M). When MS medium containing 2,4-d (5M) was used for seedling pretreatment and for tTCLs culture, somatic embryos were observed 2 weeks earlier, i.e. after 7 weeks of culture. On the tTCLs from seedlings pretreated with 2,4-d (5M) combined with benzyladenine and zeatin at 0.1 M (BZ), somatic embryos were observed after 6 weeks of culture and the percentage of embryogenesis was higher (62%) than when 2,4-d was used alone for pretreatment (40%). Similar results were also obtained from pretreatment with combinations of 2,4-d (5M) and thidiazuron (TDZ) (0.01, 0.1M). When a combination of 2,4-d (5M) and BZ (0.1M) was used both for seedling pretreatment and for tTCLs culture, both somatic embryos and shoots were observed after only 3 weeks. As the concentration of BZ increased, the percentage of somatic embryogenesis decreased but the percentage of organogenesis increased. Similar responses were obtained with a combination of 2,4-d (5M) and TDZ (0.01M). On the medium containing both NAA (0.3M) and BZ (1M), globular- and heart- stage embryos developed after 4 weeks of culture into cotyledonary-staged embryos which remained dormant after a short elongation of the embryo axis. The importance of seedling pretreatment by growth substances in enhancing somatic embryogenesis is reported.Abbreviations BA 6-benzyladenine - BZ combination of BA and zeatin - 2,4-d 2,4-dichlorophenoxyacetic acid - MS medium Murashige and Skoog basal medium - NAA a-naphthaleneacetic acid - TDZ thidiazuron - tTCLs transverse thin cell layers - TCL longitudinal thin cell layer     

A novel technique for the study of bacterial cell mechanical properties

A micromanipulation method is described for measuring the bursting forces of bacteria and relating them to cell size. At a compression speed of 6.2 m s^–1, bursting forces of three samples of rapidly growing Staphylococcus epidermis from a batch culture varied from 3 to 34 N with an average value of 13.8 N (standard error 0.8 N). Escherichia coli grown in continuous culture at a specific growth rate of 0.5 h^–1 had bursting forces varying from 1 to 9 N with an average value of 3.6 N (standard error 0.4 N). In squeeze-hold experiments, force relaxation was observed, which was attributed to water loss from the cells, or viscoelasticity, or both. At high compression speed, such as 6.2 m s^–1, this relaxation could be neglected. Micromanipulation strength measurements might be used in studies of cell mechanical disruption and of the dependence of cell strength on cell physiology.     

Regeneration of plants from callus and embryos of ‘Royal’ apricot

Immature embryos of apricot (Prunus armeniaca L.) cv. Royal with a PF index of 25–100 were used to regenerate plants in vitro using two methods. In the first case, callus was initiated on MS medium with 4.5 M 2, 4-D plus 0.44 M BA and regeneration of shoots from the callus occurred on MS medium with 4.4 M BA plus 1.0 M 2, 4-D. In the second case, adventitious buds were directly regenerated from the cotyledons on MS medium with 4.4 M BA plus 1.0 M 2, 4-D.Abbreviations BA 6-benzyladenine - IBA dole-3-butyric acid - NAA -naphthylacetic acid - 2, 4-D 2, 4-dichlorophenoxyacetic acid - PF (embryo length/seed length) x 100     

Isolation and culture of Lens culinaris Medik. cv. Eston epicotyl protoplasts to calli

Protoplasts of Lens culinaris Medik. cv. Eston were isolated from epicotyl tissues of seedlings grown on Murashige & Skoog basal medium. For isolating the protoplasts, epicotyl tissues were digested for 12–14 h at 25°C in an isolation mixture (pH 5.7) containing 1% Cellulase RS, 0.5% Driselase, 0.25% Pectolyase Y23, 0.2M calcium chloride, 10 mM mannitol and 10 mM MES. Protoplasts were purified by flotation over 20% sucrose and washed with 0.2 M calcium chloride solution supplemented with 10 mM mannitol. Purified protoplasts were cultured at a density of 10⁵ ml^-1 in agarose (Seaplaque, 0.6%) blocks which were suspended in an identical but liquid KM8P culture medium lacking amino acids, ammonium nitrate, and coconut water but containing 0.35 M glucose and a growth regulator complement of either 2.2 M 2,4-dichlorophenoxyacetic acid (2,4-D), 2.7 M naphthaleneacetic acid (NAA), 2.3 M N-(2-furanylmethyl)-1H-purine-6-amine (kinetin), 2.2 M benzylamino purine (BAP), 2.3 M 2-methyl-4-(1H-purine-6-ylamino)-2-buten-1-ol (zeatin), and 1.4 M gibberellic acid (GA₃), or 5.4 M NAA and 2.2 M each of 2,4-D and BAP. The osmotic potential of the liquid culture medium was gradually reduced over a period of 3 weeks by replacing the spent medium with a fresh medium containing 0.25, 0.1 and 0 M glucose at weekly intervals. About 6% of the dividing protoplasts developed into cell colonies after 3 weeks of culture at 25°C in diffuse light (10 E m^-2s^-1). In 35–42 days the microcolonies were about 1 mm in diameter and developed into calli on transfer to agar-solidified B5 medium supplemented with growth regulators used in the protoplast culture medium and 5 mM glutamine. Attempts to regenerate plants from protoplast-derived calli have so far been unsuccessful.Department of Applied Microbiology and Food Science, University of Saskatchewan     

Regeneration from long-term cell suspension cultures of tepary bean (Phaseolus acutifolius)

Plant regeneration has been achieved from long-term cell suspension cultures established from leaf derived callus of tepary bean (Phaseolus acutifolius). The proportion of densely cytoplasmic cells in suspension culture increased when cultured in the L-6 medium with 54 M NAA and 2 M KN. Filtration of the cells at each of five consecutive subcultures resulted in the isolation of a plant regenerating cell line (TB 686), which is being maintained in L-6 medium with 4.5 M 2,4-D and 2.3 M zeatin. Differentiated green cell aggregates were obtained when cells from maintenance medium were transferred to the same medium with 10 M BA. Embryo-like structures developed from these aggregates on L-6 medium with 2.3 M zeatin, 0.69 M GA₃ and 1.5 M NAA. Plantlets regenerated from these structures when they were cultured on L-6 medium with 7.0 M NAA and 1.0 M KN. Plant regeneration from the cell line remained relatively constant for 270 days. Regenerated plants were grown to maturity in the greenhouse.Abbreviations BA Benzyladenine - 2,4-D 2,4-dichlorophenoxyacetic acid - GA₃ Gibberellic acid - IPA Isopentenyladenine - KN Kinetin - NAA Naphthaleneacetic acid - AA Amino acid medium (Toriyama and Hinita, 1985) The research was sponsored by United States Agency for International Development, Washington D.C., Cooperative Agreement DAN-4137-A-00-4053-00     

The Sertoli cell of the water buffalo (Bubalus bubalis) during the spermatogenic cycle

Summary Ultrastructural features and morphometric evaluations of buffalo Sertoli cells are reported for the six phases of the spermatogenic cycle. The phases of the tubular seminiferous epithelium are identified according to characteristic cellular associations with completed spermiation as demarcation between two cycles. Average tubular diameter (245 m) and epithelial height (61 m) do not vary significantly during the cycle. The relative Sertoli cell volume in the seminiferous epithelium varies between 30% (phase 4) and 39% (phase 8). The calculated volume of a single Sertoli cell increases from a nadir of 7118 m³ in phase 3 abruptly to a maximum of 8968 m³ in phase 4 and is then gradually reduced during the following phases. The Sertoli cell surface area shows a similar trend: it amounts to 11105 m² in phase 3 and to 14260 m² in phase 4. The contact area of the Sertoli cell with adjacent cells and structures is subject to characteristic changes; from the expansion of basal Sertoli-Sertoli contacts it is concluded that the blood-testis barrier in the buffalo is particularly tight during phases 8, 1 and 2. The irregularly contoured nucleus contains a vesicular nucleolus, has a calculated volume from 465 m³ to 543 m³ and occupies 5 to 7% of the cell. Volume percentages of mitochondria (4%), Golgi apparatus and lysosomal bodies are rather constant during the cycle. Whorls and orderly arranged aggregates of the smooth endoplasmic reticulum occur in basal location as well as in close association with elongating spermatids. Smooth ER is the organelle that exhibits the most prominent changes during the Sertoli cell cycle: it occupies 5.79% in phase 3 and 20.9% in phase 4 of the total cellular volume. Phagocytosis of residual bodies is insignificant in this species and a lipid cycle is absent in buffalo Sertoli cells.     

Phenylacetic acid-induced somatic embryogenesis in cultured hypocotyl explants of geranium (Pelargonium x hortorum Bailey)

Summary The effect of a non-indole compound, phenylacetic acid (PAA), on the induction of somatic embryogenesis in tissue cultures of geranium (Pelargonium x hortorum Bailey cv. Scarlet Orbit Improved) was investigated. Hypocotyl explants derived from young, dark-grown seedlings were cultured on Murashige and Skoog (1962) medium (MS) supplemented with PAA or IAA (0.01–120 M) alone or in combination with BAP (8 M). Somatic embryogenesis was induced by both PAA and IAA at 0.01–20 M with 8 M BAP, however, the optima differed considerably for the two compounds. Maximal activity of IAA for somatic embryogenesis was found at 0.1–2.5 M, whereas PAA gave best results at 10 and 20 M under identical culture conditions. Higher concentrations (30–120 M) of IAA or PAA in the medium induced callusing in the explants, but the callus was neither embryogenic nor morphogenic.Abbreviations BAP N⁶-benzylaminopurine - IAA indole-3-acetic acid - MS Murashige and Skoog (1962) - PAA phenylacetic acid     

In vitro binding of riboflavin to subcellular particles from maize coleoptiles and Cucurbita hypocotyls

Saturable and reversible in vitro binding of [¹⁴C]riboflavin was found to occur on subcellular, sedimentable particles from maize coleoptiles and Cucurbita hypocotyls. The K_D was ca. 6 M, the pH optimum was near 6.0, and the number of binding sites amounted to 0.1–0.5 M on a fresh-weight basis. When the reducing agent dithionite was present, riboflavin binding increased-the K_D was 2.5 M, and the pH optimum above 8.0. The binding was specific: flavin mononucleotide (FMN) and flavin adenosine-dinucleotide (FAD) bound less tightly to these sites than riboflavin and another major soluble flavin, the previously described riboflavin-analog FX, occurring in grass coleoptiles. These flavin-binding sites were localized on vesicles derived from plasmalemma and endoplasmic reticulum by analyzing sucrose and metrizamide density gradients and marker enzymes.Abbreviations CCO cytochrome-c oxidase - CCR NADH-cytochrome-c oxidoreductase - ER endoplasmic reticulum - FAD flavin-adenosinedinucleotide - FMN flavin mononucleotide - MOPS N-morpholino-3-propansulfonic acid - NADH reduced -nicotinamide dinucleotide - nKP n thousand times g pellet - NPA l-naphthylphthalamic acid - PM plasma membrane, plasmalemma - RBF riboflavin - IAA indoleacetic acid - BA benzoic acid     

In vitro propagation of Asparagus maritimus – A rare Mediterranean salt-resistant species

Asparagus maritimus L. Miller is a rare species growing of the Mediterranean region and is morphologically similar to A. officinalis. In order to establish an efficient in vitro propagation protocol, explants were excised from spear segments and cultured on Murashige and Skoog (1962) medium containing 3% sucrose and various concentrations of growth regulators. The best shoot initiation (3–4 per explant) was achieved on a medium containing 0.88 M N⁶-benzyladenine (BA), 0.93 M kinetin, 1.07 M -naphthaleneacetic acid (NAA) and 3.90 M ancymidol. Shoot initiation could also be achieved without ancymidol but the shoots were thinner and longer. A very high shoot multiplication rate was achieved on media supplemented with 3% sucrose, 1.07 M NAA, 0.93 M kinetin, 0.44 M BA and various concentrations of ancymidol. The lowest concentration of ancymidol (0.39 M) significantly promoted the highest shoot multiplication rate (11.9 shoots/crown). For root formation, media were supplemented with 6% sucrose, 1.07 M NAA and various concentrations of ancymidol. Rooting frequency increased with higher ancymidol concentration up to 5.07 M (82.0% rooting). The number of ex vitro shoots formed was strongly correlated (r=0.66) with the length of roots formed in vitro, which was the highest at a 1.95 M ancymidol.     

Purification and characterization of flavone-specific β-glucuronidase from callus cultures of Scutellaria baicalensis Georgi

-Glucuronidase from callus cultures of Scutellaria baicalensis Georgi was purified to apparent homogeneity by fractionated ammonium-sulfate precipitation and chromatography on diethylaminoethyl-cellulose, hydroxylapatite and baicalin-conjugated Sepharose 6B. A 650-fold purification was obtained by this purification system. When subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis the purified protein migrated as a single band with a molecular mass of 55 kDa. We determined that the native enzyme has a molecular mass of 230 kDa using gel-filtration chromatography. These results suggested that the enzyme exists as a homotetramer composed of four identical 55-kDa subunits. The enzyme showed a broad pH optimum between 7.0 and 8.0. The K _m values were 9 M, 10 M, 30 M and 40 M for luteolin 3 -O--d-glucuronide, baicalin, wogonin 7-O--d-glucoronide and oroxlin 7-O--d-glucuronide, respectively. The enzyme was most active with flavone 7-O--d-glucuronides.Abbreviations BA N⁶-benzyladenine - 2,4-D 2,4-dichlorophenoxyacetic acid - pI isoelectric point - R _t retention time     

Plant regeneration from embryo-derived callus of oil palm – the effect of exogenous polyamines

Regeneration in oil palm was achieved through somatic embryogenesis/organogenesis from embryo-derived callus. Callus was induced from mature embryos of the cross 281 (D)×18 (P) on modified MS medium supplemented with 2,4-D (113.12 M) and 2-iP (14.76 M). The embryogenic calluses obtained were transferred to Blaydes medium supplemented with 2,4-D (0.045 M) and one of the following growth regulators: TDZ (4.54 M), zeatin riboside (2.85 M), putrescine (1 mM) and spermine (100 M). Secondary somatic embryogenesis was found to occur in media supplemented with polyamines. The efficiency of formation of somatic embryos, secondary somatic embryos and shoot meristemoids were significantly higher in putrescine containing medium. Histological studies were also undertaken.     

Somatic embryogenesis in callus cultures of Rosa hybrida L. cv. Landora

Callus was initiated from immature leaf and stem segments of rose (Rosa hybrida cv. Landora) and subcultured every four weeks on a basal medium of half-strength Murashige & Skoog (1962) salts plus 30 g l^-1 sucrose (1/2 MS) and supplemented with 2.2 M BA, 5.4 M NAA and 2.2–9.0 M 2,4-D. Embryogenic callus and subsequently somatic embryos were obtained from 8-week-old callus culture on 1/2 MS+2.2 M BA+0.05 M NAA+0.3 M GA₃+200–800 mg l^-1 L-proline. Long-term cultures were established and maintained for up to 16 months by repeated subculture of embryogenic callus on L-proline deficient medium. About 12% of cotyledonary stage embryos taken from cultures cold-stored at 8±1°C for 4 days germinated on 1/2 MS+2.2 M BA+0.3 M GA₃+24.7 M adenine sulphate.Abbreviations BA benzyladenine - NAA -naphthaleneacetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - GA₃ gibberellic acid     

Axillary shoot induction and plant regeneration in Plantago ovata Forssk

Axillary shoot induction and plant regeneration were obtained in Plantago ovata. The optimum medium for inducing axillary shoots was Murashige & Skoog (MS) medium [5] supplemented with 4.6 M kinetin and 0.05 M NAA. Rooting of shoots was best on half-strength MS medium containing 5.0 M IBA and 0.05 M kinetin. The regenerated plants were similar to the control plants in karyotypic and phenotypic details.     

Effects of specific inhibitors of cellular functions on sulfur mustard-induced cell death

This study was conducted to determine whether inhibitors of normal cellular functions can reduce cytotoxicity induced by sulfur mustard (HD). The compounds examined include inhibitors ofpoly(ADP-ribose) polymerase (PADPRP), inhibitors of mono(ADP-ribose) transferase (MADPRT), inhibitors of lipidperoxidation, and an inhibitor ofprotein synthesis. To determine the effects of these compounds on HD-induced cell death, human lymphocyte preparations were treated with known concentrations (0.1 M to 1000 M) of an inhibitor and exposed to an estimated 87% effect concentration (EC₈₇) of HD (170 M) for loss in cell viability. Cell viability was determined at 24–26hr post-exposure to HD using a dye (propidium iodide) exclusion assay and a flow cytometer. All of the selected PADPRP inhibitors were found to be effective at reducing the cytotoxic effects of HD. These inhibitors were rank-ordered based on the concentration that gives 50% (EC₅₀) reduction ofHD-induced cell death.A signijicant correlation (r=0.94) was observed between the compounds' ability to inhibit PADPRP and the compounds' ability to reduce HD- induced cell death, suggesting that PADPRP plays a role in HD-induced cell death. Inhibitors of MADPRT, lipid peroxidation, and protein synthesis were not effective at reducing HD-induced cell death.Abbreviations ATP adenosine triphosphate - DNA deoxyribonucleic acid - EC₅₀ concentration which gives 50% of maximum effect - GSH glutathione - HD sulfur mustard (,-dichloroethyl sulfide) - HEPA high efficiency particulate adsorbing - HEGA high efficiency gas adsorbing - IC₅₀ concentration that inhibits 50% of enzyme activity - MADPRT mono(ADP-ribose) transferase - NAD nicotinamide adenine dinucleotide - PADPRP poly(ADP-ribose) polymerase     

Hormone-induced protection of sunflower photosynthetic apparatus against copper toxicity

The effects of excess Cu as affected by the application of exogenous hormones (gibberellic acid - GA₃ and indole-3-acetic acid - IAA) with respect to sunflower (Helianthus annuus L.) growth, physiology, and metabolism were studied. Application of 100 M IAA lessened the toxic effects of 80 M Cu in roots indicating greater root length and root hair formation, while addition of 100 M GA₃ ameliorated the toxic effect mainly to the shoot. The content of photosynthetic pigments significantly declined under Cu stress, whereas application of hormones led to a substantial preservation of chlorophylls and carotenoids. Under Cu stress, the rate constant of energy trapping by photosystem 2 (PS2) reaction centres (RCs) was reduced as a result of physical dissociation of the light-harvesting complex (LHC) from PS2 core, while application of IAA and especially GA₃ resulted in stability of the LHC of PS2 RCs. The drop in net photosynthetic (PN) and transpiration (E) rates with preserved or slightly reduced variable to maximum fluorescence ratio (F_v/F_m) in the presence of 80 M Cu could be explained by a possible inhibition of the enzymatic processes in the Calvin cycle. Application of 100 M IAA and 100 M GA₃ lessened Cu effects mainly on P _N. Water use efficiency was also improved under hormone exposure.     

Different effects of inorganic and triethyl lead on growth and ultrastructure of lily pollen tubes

Summary Pollen tubes ofLilium longiflorum growingin vitro were treated for 1 h with inorganic lead (Pb) and with triethyl lead (TriEL) and studied by light and electron microscopy. Pb was considerably more toxic in relation to inhibition of pollen tube growth (EC₅₀=6 M Pb) than was TriEL (EC₅₀=60 M TriEL). On the other hand, at almost the entire concentration range tested (25-500 M) TriEL caused aberrant tubes and tube swellings. Pb did not cause tube swellings, even at highly growth-impairing concentrations. Pb (60 M) predominantly affected the ultrastructure of the growing cell walls without impairing the distribution of the cell organelles in the tube tips. In contrast, 50 and 100 M TriEL did not visibly influence cell wall ultrastructure but it severely damaged dictyosomes; 100 M TriEL also disturbed the original order of cell organelles in the tube tips. Cortical microtubules were selectively and completely destructed by TriEL at concentrations (50 M) where no effect on polar organization of the tube tips occurred but they remained unimpaired by 60 M Pb, indicating selective and effective interaction of TriEL with these cell organelles.Abbreviations EC⁵⁰ effective lead concentration causing 50% inhibition of pollen tube growth - MTs microtubules - Pb inorganic lead - TriAL trialkyl lead - TriEL triethyl lead     

Transient state characteristics of adaptation to changes in light conditions for the cyanobacterium Oscillatoria agardhii

Transitions in growth irradiance level from 92 to 7 Em^-2 s^-1 and vice versa caused changes in the pigment contents and photosynthesis of Oscillatoria agardhii. The changes in chlorophyll a and C-phycocyanin contents during the transition from high to low irradiance (HL) were reflected in photosynthetic parameters. In the LH transition light utilization efficiencies of the cells changed faster than pigment contents. This appeared to be related to the lowering of light utilization efficiencies of photosynthesis. As a possible explanation it was hypothesized that excess photosynthate production led to feed back inhibition of photosynthesis. Time-scales of changes in the maximal rate of O₂ evolution were discussed as changes in the number of reaction centers of photosystem II in relation to photosynthetic electron transport. Parameters that were subject to change during irradiance transitions obeyed first order kinetics, but hysteresis occurred when comparing HL with LH transients. Interpretation of first order kinetic analysis was discussed in terms of adaptive response vs changes in growth rate.Non-standard abbreviations Chla chlorophyll a - CPC C-phycocyanin - PS II photosystem II - PS I photosystem I - RC II reaction center of photosystem II - P photosynthetic O₂-evolution - I irradiance, Em^-2 s^-1 - light utilization efficiency of cells, mmol O₂·mg dry wt^-1·h^-1/Em^-2 s^-1 - light utilization efficiency of photosynthetic apparatus, mol O₂·mol Chla ^-1·h^-1/Em^-2 s^-1 - P_max maximal rate of O₂ evolution by cells, mol O₂·mg dry wt^-1·h^-1 - P_max maximal rate of O₂ evolution by photosynthetic apparatus, mol O₂·mol·Chla ^-1·h^-1 - LL low light, E m^-2 s^-1 - HL high light, E m^-2 s^-1 - LH low to high light transition - HL high to low light transition - k specific rate of adaptation, h^-1 - specific growth rate, h^-1 - Q pool size of cell constituent, mol·mg dry wt^-1 - q net synthesis rate of cell constituent, mol·mg dry wt^-1·h^-1     

Tissue culture ofKarwinskia humboldtiana—a plant producing toxins with antitumoural effects

Isolated embryos ofKarwinskia humboldtiana were cultured in vitro. The growth of embryos and development to plantlets on woody plant medium supplemented with indole-3-acetic acid 6.10^-2 mol l^–1, gibberellic acid (GA₃) 3.10^-2 mol l^–1, and 6-benzylaminopurine (BA) 2 mol l^–1 was obtained. Multiplication of shoots and rooting of excised shoots has been achieved. Callus formation on modified Murashige-Skoog medium supplemented with 1-naphthaleneacetic acid 10 mol l^–1, GA₃ 14 mol l^–1, and kinetin 5 mol l^–1 on hypocotyls, or on root cultures on medium supplemented with 2.4-dichlorophenoxyacetic acid 10 mol l^–1 and BA 10 mol l^–1 was induced.Abbreviations BA 6-benzylaminopurine - 2,4-d 2,4-dichlorophenoxyacetic acid - GA₃ gibberellic acid - IAA indole-3-acetic acid - NAA 1-naphthaleneacetic acid - TEM transmission electron microscopy