In vitro plantlet regeneration of<Emphasis Type="Italic"> Schinopsis balansae</Emphasis> (Anacardiaceae)

A protocol for the in vitro regeneration of rooted plants from nodal single bud segments of 10-year-old Schinopsis balansae trees was developed. Nodal segments were harvested from actively growing shoots of plants grown from seeds and maintained in pots under greenhouse conditions, and from epicormic shoots obtained by forced flushing of branches. Culture of nodal segments on nutrient medium containing the mineral salts and vitamins of Murashige and Skoog medium at 1/4 strength (1/4 MS), supplemented with 100 mg l^–1 ascorbic acid, 3% sucrose, and 5–15 M 6-benzyladenine resulted in regeneration of multiple shoots. Rooting of regenerated shoots was observed in 1/4 MS medium with vermiculite as the substrate and supplemented with 7.5 M indolbutyric acid.     

Rapid clonal multiplication of Lavandula viridis L'Hér through in vitro axillary shoot proliferation

In vitro clonal propagation of native Mediterranean Lavandula viridis was obtained from a mature field-grown plant. Single node explants were successfully established on Murashige and Skoog medium supplemented with 0.44 M of 6-benzyladenine. The highest multiplication rate (11.69 shoots/node) was obtained with 0.67 M 6-benzyladenine in Murashige and Skoog medium with macronutrients at half-strength. Shoots were easily rooted on Gresshoff and Doy medium. Increasing sucrose concentration from 58.4 to 87.6 mM resulted in a significant increase in rooting frequency. Eighty per cent of plantlets were successfully acclimatised to ex vitro conditions, exhibiting a normal development.     

Micropropagation of Limonium cavanillesii Erben, a threatened statice, from inflorescence stems

Micropropagation of Limonium cavanillesii Erben, a threatened and endemic statice species from Valencia Community (Eastern Spain), was successfully achieved using inflorescence stem pieces as initial explants. Segments 20 mm long from basal parts of immature inflorescences and with axillary buds were cut, sterilised and established in vitro. Shoots obtained from indifferentiated buds were sectioned and then transferred to Murashige and Skoog (MS) medium with 2 mg l^–1 kinetin to provide a plant stock.Shoot multiplication was achieved on MS medium with different cytokinins. The best results for shoot formation were obtained with 2–5 mg l^–1kinetin, 5 mg l^–1 6---dimethylallylaminopurine or 0.1 mg l^–1 6-benzylaminopurine, without significant differences between them. High shoot rooting (80–85%) was obtained within four weeks with indolebutyric acid or indoleacetic acid (0.1 or 0.5 mg l^–1), and also on medium without plant growth regulators. Plant survival to hardened greenhouse conditions was 90% four weeks after plantlet removal from in vitro conditions.This protocol for micropropagation of Limonium cavanillesii is very useful for conservation purposes of endangered statice species, because by using inflorescence stem as initial material it is easier to establish aseptic cultures while preserving the mother plant.     

In vitro rapid regeneration of plantlets from nodal explants of Mucuna pruriens– a valuable medicinal plant

A rapid and efficient protocol for the large‐scale propagation of a potential medicinal plant, Mucuna pruriens, through in vitro culture of nodal segment explants obtained from 15‐day‐old aseptic seedlings is described. Of the three different cytokinins, 6‐benzyladenine (BA), kinetin (Kin) and 2‐isopentenyl adenine (2‐iP) evaluated as supplements to Murashige and Skoog (MS) medium, BA at an optimal concentration of 5.0 μM was effective in inducing multiple shoots. Strength of the basal media also influenced the efficiency of shoot regeneration. The frequency of shoot regeneration tended to increase when the salt concentration in the basal media was reduced. Highest number of multiple shoots (23.3) and maximum average length (5.6 cm) were standardised on half‐strength MS medium supplemented with 5.0 μM BA along with 0.5 μM α‐naphthalene acetic acid (NAA) at pH 5.8. Rooting was best induced in shoots excised from proliferated shoot cultures on MS medium augmented with an optimal concentration of 1.0 μM indole‐3‐butyric acid (IBA). The in vitro‐raised plantlets with well‐developed shoots and roots were successfully established in earthen pots containing garden soil and were grown in greenhouse with 90% survival rate. The results of this study provide the first report on in vitro plant regeneration of M. pruriens.     

影响油菜子叶外植体不定芽高频率再生的因素

采用７个甘蓝型和２个白菜型油菜品种研究了影响子叶外植体芽再生的一些因素和不同基因型的子叶外植体的离子体培养反应和芽再生能力。结果表明,苗龄４ｄ的幼苗叶子含２,４－Ｄ０．５－１．２ｍｇ／Ｌ和６－ＢＡ０．２ｍｇ／Ｌ的培养基培养２或６ｄ后再转到分化培养基上培养,芽再生率３４％－４６％。     

Effects of auxin on the spatial distribution of cell division and xylogenesis in lettuce pith explants

Summary Cylinders of pith parenchyma were tissue-cultured with their opposite ends on media which differed only in content of the morphogens auxin (IAA), sucrose, or zeatin. A range of concentrations of each of these morphogens applied at one end (none at the other end) resulted in distribution patterns of cell division and xylogenesis that were attributable to interaction between inductive levels and morphogen mobility. Auxin was crucial for tracheary patterns: large tracheary elements formed by direct differentiation of pith cells near the auxin source, smaller but still roughly isodiametric tracheary elements formed after cell division, and tracheary strands developed where, presumably, auxin transport had become polarized and then canalized. Xylogenesis was confined to regions within millimeters of the auxin source, and [¹⁴C]IAA studies showed a steep logarithmic concentration gradient along the cylinder. Patterns of tracheary strands and rings revealed that the pith explants retained some polarity from the stem from which they had been excised. However, the direction of flow of applied auxin was more effective than original polarity in controlling the orientation of tracheary strands and their constituent tracheary elements. It seems that, in tissues with little or no polarity, diffusive flow of auxin gradually induces polar flow in the same direction, together with an associated bioelectric current, and that this orients the cortical microtubules that in turn determine the orientations of cell elongation and of the secondary wall banding in tracheary elements.Abbreviations IAA indoleacetic acid - NAA naphthaleneacetic acid - TIBA triiodobenzoic acid Dedicated to the memory of Professor John G. Torrey     

Regeneration of niger (Guizotia abyssinica Cass.) CV Sahyadri from seedling explants

Root, hypocotyl and cotyledonary explants of niger (Guizotia abyssinica Cass) CV. Sahyadri were aseptically cultured on Murashige and Skoog's basal medium (MS) containing BAP and kinetin. Multiple shoot regeneration was induced from hypocotyl and cotyledonary explants while root explants produced only callus on MS medium supplemented with BAP. BAP (1 mg l^-1) was optimum for shoot regeneration. Regenerated shoots were transferred to MS medium without auxins, with auxins and with increasing concentrations of sucrose for rooting. Complete plantlets were obtained in all cases; however, 0.5 mg l^-1 NAA was the best for induction of roots. Ninety-seven per cent of the plantlets survived and completed their life cycle when transferred to natural conditions.Abbreviations BAP 6-benzylamino purine - NAA -naphthaleneacetic acid - IAA indole-3-acetic acid - IBA indole-3-butyric acid     

The effects of cadmium exposure on the cytology and function of primary cultures from rainbow trout

Cultured epidermal cells from explants of skin of rainbow trout were used to study the cytological and functional changes following sublethal exposure to cadmium stress. The aim was to develop diagnostic markers for ecotoxicology. Cultures were exposed to the pollutant for 48 h. Cell structural and cytological changes were established by light and electron microscopy. Metabolic alterations were detected by immunohistochemistry. The relation between the initiation of cellular alterations and cadmium concentrations was compared in cultures exposed in commercially-available serum-free and serum-containing medium. The expression of stress proteins (metallothionein and heat shock protein) was also studied. Rainbow trout epithelial cells exposed to cadmium showed typical morphological changes indicative of cell death by apoptosis. Sublethal exposure also resulted in cellular metabolic disturbances with increased deposits of glycogen. Increased melanization was also observed. These changes appeared at lower concentrations of cadmium when cells were exposed in serum-free media than in serum-containing media. Cadmium induced the expression of heat shock proteins but not of metallothioneins. The results broadly confirm in vivo findings for cadmium toxicity and suggest that this in vitro technique may have applications in aquatic toxicology. © 1998 John Wiley & Sons, Ltd.     

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) inhibition of coronary vasculogenesis is mediated, in part, by reduced responsiveness to endogenous angiogenic stimuli, including vascular endothelial growth factor A (VEGF-A)

BACKGROUND: 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) exposure prior to chick embryo incubation (GD 0) induces dilated cardiomyopathy, and reduces myocardial hypoxia, vascular endothelial growth factor A (VEGF-A) expression, and coronary vascularization. We investigated whether reduced coronary vascularization 1) occurs in the absence of changes in cardiac morphology and 2) is associated with altered secretion of VEGF-A and/or an antivasculogenic factor. METHODS: Chicken eggs were treated with control (corn oil) or TCDD (0.075-0.3 pmol of TCDD/gm) on GD 5. In vivo cardiac morphology and artery number were determined on GD 10, while in vitro vascular outgrowth and VEGF-A secretion were determined from cardiac explants on GD 6. Effects of recombinant VEGF-A (rcVEGF-A), soluble flt-1 (sFlt-1) receptor plus rcVEGF-A, and control conditioned media were assessed in TCDD explants, while effects of TCDD-conditioned media was assessed in control explants. RESULTS: TCDD reduced coronary artery number in vivo by 53 +/- 8% and induced a dose-related reduction in tube outgrowth in vitro, but had no effect on cardiac morphology. All TCDD doses reduced explant VEGF-A secretion equally (43 +/- 3%), compared to control. sFlt-1 blocked outgrowth in control cultures and blocked rcVEGF-A-mediated rescue of outgrowth in TCDD explants. Control conditioned media partially rescued outgrowth from TCDD explants, while conditioned media from TCDD explants had no effect on controls. CONCLUSIONS: TCDD inhibition of coronary vascularization can occur in the absence of changes in cardiac morphology and is associated with reduced VEGF-A secretion but not an antivasculogenic factor. Since control media only partly rescues TCDD's inhibitory effect, we suggest that TCDD-exposed endothelial cells are less responsive to vasculogenic stimuli.     

Enhanced bud regeneration in aspen (Populus tremula L.) roots cultured in liquid media

 The regeneration potential of excised aspen (Populus tremula L.) roots cultivated in liquid medium, as affected by plant growth regulators and by the position of the isolated root explant on the main root, was investigated. The effect of various levels of benzyladenine (BA) and thidiazuron (TDZ) on bud regeneration in root explants was studied. TDZ in the medium had a marked effect on bud development as compared with BA, inducing a tenfold increase in the number of buds regenerated from various root explants. TDZ enhanced both root and root-borne shoot biomass production but reduced further shoot development and elongation. The position of the isolated root sections on the main root affected regeneration, the proximal sections further away from the root tip producing the highest number of buds per explant in both BA and TDZ treatments. Buds regenerated in close proximity to the site of lateral roots in BA-treated roots, while in TDZ-treated root sections, the buds formed all over the root regardless of the presence of lateral roots. The buds developed from inner cortical and sub-epidermal cell layers, disrupting the epidermis and the inner layers. Root biomass production and growth was greatly enhanced in well-aerated bioreactor culture in the presence of 4.5×10^–2 μM TDZ. A high number of the root-borne shoots could be rooted and converted to plantlets. However, while shoots regenerated in a medium with BA rooted well in a growth regulator-free medium, shoots formed in a medium with TDZ required auxin for rooting. Roots cultured in the presence of ancymidol, a gibberellin biosynthesis inhibitor, regenerated non-hyperhydric bud clusters and hyperhydric shoots. These were separated mechanically, subcultured to growth and rooting medium and transplanted ex vitro resulting in phenotypically true-to-type plantlets. The potential of liquid cultures for aspen shoot biomass production from roots is discussed. Received: 24 January 2000 / Revision received: 6 March 2000 / Accepted: 7 March 2000