<Emphasis Type="Italic">In vitro</Emphasis> regeneration,flower and plant formation from petiolar and nodal explants of culantro (<Emphasis Type="Italic">Eryngium foetidum</Emphasis> L.)

Summary A procedure for plant regeneration, flower and plant formation from petiolar and inflorescence nodal explants of culantro is discribed. Leaf petioles were excised from young leaves of non-flowering plants while nodal explants were excised from the inflorescence. Explants were cultured in Murashige and Skoog (MS) medium alone or supplemented with 0.5μM naphthaleneacetic acid (NAA) and 0.9, 1.8, 4.5, or 9 μM thidiazuron (TDZ). All explants produced multiple shoots. In addition, nodal explants formed flowers. Shoot number, flower number and shoot length were influenced by TDZ and NAA. Rooted shoots from both types of explants were transferred to soil where plants were successfully established.     

Transition from absolute to quantitative short-day control in Nicotiana tabacum cv. Maryland Mammoth

The response in vitro of thin cell layers, excised from different stem regions of Nicotiana tabacum cv. Maryland Mammoth plants at various developmental stages, was studied under different photoperiodic treatments. The aim was to determine at which stage of plant development, and in which region of the stem, the absolute short-day requirement, indispensable for the induction of the flowering process in this genotype, becomes quantitative and whether it remains short-day. The explants were cultured on a medium suitable for flower neoformation, and were exposed for 30 days to the following treatments: continuous darkness, 8 h light/16 h dark per day, 16 h light/8 h dark per day, and continuous light. The first flowers on explants were observed from plants that were still in the vegetative state, but whose apex showed an accelerated production of axillary vegetative buds, as observed histologically. These explants were excised from the first 10 internodes below the first node with a leaf ≥ 5 cm in length (apical site), and produced flowers only under short-day treatment. When the apical dome initiated the organization of the terminal flower, the apical site explants developed flowers under both short-day and long-day treatments. At the same stage, explants from the 15th to the 20th internode below the first leaf ≥ 5 cm in length also formed flowers, but only under short-day. When the plant showed a complete inflorescence, flowers were also present on explants from the most basal stem internodes and from the inflorescence branches. At this stage, flower neoformation occurred under all treatments; however, under short-day the number of explants showing flowers not associated with vegetative buds on the same sample greatly exceeded that observed under other treatments, as did the mean number of flowers per explant (except the basal regions). In conclusion, in the post-inductive phases of the flowering process, the photoperiodic requirement of this genotype is always short-day. The superficial tissues of the stem require either absolute or quantitative short-day treatment, depending on their position on the stem and the stage of evolution of the flowering process in the terminal apex.     

Consequences of the Elimination of Old Leaves upon Spring Phenological Events and the New Leaves Nutrient Concentration in a Wintergreen Woody Species in the Southern Hemisphere

Previous studies analyzed the importance of old leaves conservancy for wintergreen species plant growth only after early spring old leaves elimination. However, carbon and nutrient resources for growth could have already been translocated from old leaves to shoots during autumn. In this work, the effect of old leaves absence on the leaf mass per area (LMA, g m⁻²) and nutrient concentration of new spring leaves, shoot growth, and flowering was studied in Aristotelia chilensis, an Andean Patagonic woody wintergreen species of Argentina. Plants were studied after autumn defoliation (AD) or late winter defoliation (WD) and results were compared to those of undamaged control plants (CO). The new leaves LMA and mineral nutrient (N, P, K, and Mg) concentration values did not decrease in AD or WD compared to CO plants. Conversely, CO plants showed higher flowering intensity and shoot lengthening compared to AD or WD plants. There were not remarkable differences regarding the defoliation time, though non-flowering shoots grew in a lesser degree than the flowering shoots in WD plants. It was concluded that A. chilensis old leaves cohort is an important source to shoot growth and flowering but their absence does not affect the new leaves structure or nutritional status from early spring in either AD or in WD plants. New leaves formation probably is guaranteed by resources (carbon and nutrients) previously stored in stems or even in the buds containing the preformed leaves since March, by the end of summer. Provided the availability of complete resources for the new leaf flush independently of the old leaves A. chilensis would restore the carbon balance as soon as possible to resume the growth of heterotrophic tissues at normal rates. Endogenous response to counterbalance the old leaves absence on non-flowering shoots was more effective when there was greater lag time between defoliation and shoot growth resume. Flowering and non-flowering shoots compete for the available resources when A. chilensis have not yet expanded leaves and shoots supporting reproductive structures were stronger sinks compared to non-flowering shoots in WD plants.     

Shoot growth and distribution pattern ofCarex kobomugi in a natural stand

To clarify the growth and distribution ofCarex kobomugi, I surveyed the shoot heights and weights of a population growing in a sand dune at Sindu-ri, Wonbuk-myeon, Taean-gun, in Chungnam Province, Korea. During the growing season, size classes, based on leaf number and shoot heights, shifted, with those in the medium class moving to higher classes. Although the frequencies of those class characters showed a normal distribution curve throughout the season, the frequencies of each class based on shoot weight were evenly distributed in all size classifications. Coefficients of variation were 0.17 for leaf number, and 0.35 for leaf length and weight per plant. The maximum numbers of leaves were 8.16 ± 1.38 per plant for those that were non-flowering, but 2.66 ± 0.62 per plant for those that did flower. Non-flowering plants exhibited withered leaves by mid-September, while withering began in male plants by mid-May and by mid-July for females. At the end of the growing season, the lengths and weights of leaves from non-flowering plants were 47.8 ± 16.6 cm and 1773 ± 628 mg, respectively. When leaf order was considered, leaves increased in size along two-thirds of the ranking, then decreased. In a separate analysis, the growth ofCarex plants was compared with those ofElymus mollis in the same sample quadrats. Biomass of the former accounted for only a small portion of the total biomass per unit area (E. mollis having a dry weight of >76.4 g m^-2), but under such competition, the leaf lengths and individual plant weights nonetheless increased forCarex as well.     

Canopy development of a determinate and an indeterminate cultivar of Vicia faba L. under contrasting plant distributions and densities

Field experiments were conducted in 1987 and 1988 to quantify differences in canopy formation between an indeterminate and a determinate genotype of Vicia faba L., grown at two plant densities and three spatial distributions. The number of stems per unit area produced by determinate plants was related to the growth rate before flowering. Leaf production per stem per unit of thermal time was similar in both plant types, but twice as many leaves per stem were produced by the indeterminate cultivar. The indeterminate cultivar produced fewer and smaller leaves in the warmer and drier weather of 1988 than in 1987. The determinate genotype produced similar sizes and numbers of leaves in both years, but fewer tillers developed in 1988 than in 1987. Accordingly, leaf mass per unit ground area was greater in 1987 than in 1988 in both genotypes. Except during early flowering, relationships between leaf mass and leaf area were constant, with higher specific leaf areas in the determinate than the indeterminate genotype. Shoot dry matter partitioning into leaves was identical in both years for indeterminate plants, but differed in determinate ones.
It is concluded that canopy development is regulated through individual leaf weight and leaf number per stem in non-tillering indeterminate, and by stem numbers per unit area in tillering determinate plants.     

Field performance of micropropagated forestry species

Summary Micropropagated plants both from angiosperms (bamboo, birch, eucalyptus, tamarind, teak, willow) and gymnosperms (Douglas-fir, loblolly pine, Monterey pine, and redwood) have been established in the field. Plantlets were regenerated from juvenile explants (via adventitious or axillary buds) as well as explants from mature trees [apical and axillary buds (nodal segments)]. Plantlets regenerated from adventitious buds tend to show early maturation traits (Douglas fir, loblolly pine, Monterey pine). A population of selected clones showed superior performance and yield over seedlings derived from the same trees. Increased biomass production was obtained with plantlets derived from tissue culture ofEucalyptus spp. when compared to seedlings. No morphologic variation was observed in micropropagated plants. Plantlets derived from tissue culture grew very uniformly. Early flowering was observed with plantlets derived from tissue culture (tamarind, teak). Based on a presentation at an International Training Course on the Application of Biotechnology in Forest Trees held in Caracas, Venezuela, May 1991.     

An improved micropropagation of <Emphasis Type="Italic">Eclipta alba</Emphasis> by in vitro priming with chlorocholine chloride

An efficient method of micropropagation for Eclipta alba from young nodal axils of shoot tip explants has been developed by giving special attention to ‘priming’ in vitro plantlets in view of increasing their hardening ability after transplantation ex vitro. Among 3 cytokinins—BAP, kinetin and TDZ, BAP was found most effective in inducing and proliferating adventitious shoots. The highest frequency of responding explants (100%) and maximum number of shoots (23.0) per explant were obtained after 60 days culture on MS medium containing 8.8 μM BAP. Cent percent shoots developed roots directly from shoot base when transferred to growth regulator-free MS medium. For priming E. alba microshoots, 6.3 μM of chlorocholine chloride (CCC) was found most effective. The major changes observed in 30 days old treated shoots were, production of increased number of root, elevation of chlorophyll level in leaves and increase in plant biomass. Furthermore, arrested undesirable shoot elongation made the plants sturdier and more suitable for acclimatization. The primed micropropagated E. alba plants were healthy and survived by higher frequency (100%) in soil in comparison to the non-treated plants (84% survival).     

EFFECTS OF LEAF EXCISION ON FLOWERING OF XANTHIUM APICAL BUDS IN CULTURE UNDER INDUCTIVE AND NONINDUCTIVE PHOTOPERIODS

Apical buds of Xanthium were grown in aseptic culture under short-day cycles known to induce flowering in the intact plants or under “light-break” conditions known to prevent flowering. The total light provided in each 24-hr cycle was the same under the two photoperiods. Various numbers of leaves were excised from the apical buds. Excision of leaves did not change the response to photoperiod: even with all leaves excised the apical buds cultured under short-day conditions reached the same average floral stage as the control buds, and those under light-break conditions all remained vegetative. Fresh weight was not significantly changed by the excisions, either. However, excision of the young leaves resulted in an increase in the number of new leaves developed by the apical bud during the two-week culture period.     

What is going on with the hormonal control of flowering in plants?

Molecular genetic studies using Arabidopsis thaliana as a model system have overwhelmingly revealed many important molecular mechanisms underlying the control of various biological events, including floral induction in plants. The major genetic pathways of flowering have been characterized in-depth, and include the photoperiod, vernalization, autonomous and gibberellin pathways. In recent years, novel flowering pathways are increasingly being identified. These include age, thermosensory, sugar, stress and hormonal signals to control floral transition. Among them, hormonal control of flowering except the gibberellin pathway is not formally considered a major flowering pathway per se, due to relatively weak and often pleiotropic genetic effects, complex phenotypic variations, including some controversial ones. However, a number of recent studies have suggested that various stress signals may be mediated by hormonal regulation of flowering. In view of molecular diversity in plant kingdoms, this review begins with an assessment of photoperiodic flowering, not in A. thaliana, but in rice (Oryza sativa); rice is a staple crop for human consumption worldwide, and is a model system of short-day plants, cereals and breeding crops. The rice flowering pathway is then compared with that of A. thaliana. This review then aims to update our knowledge on hormonal control of flowering, and integrate it into the entire flowering gene network.     

Development of Agrobacterium-mediated transformation technology for mature seed-derived callus tissues of indica rice cultivar IR64

Indica rice cultivar IR64 is most recalcitrant to regenerate, which affects the transformation efficiency especially when mature seed-derived callus tissues are used as explants. Therefore, a simple, rapid and improved genetic transformation protocol has been developed for the indica rice cultivar IR64 using Agrobacterium-mediated genetic transformation. With different hormonal combination tested, the maximum callus induction was observed on MS medium supplemented with 2.5 mg/l 2,4-D and 0.15 mg/l BAP from the scutellum explants. Three weeks old scutellum derived callus explants were immersed in Agrobacterium suspension (strain LBA4404, OD600=1.0) and co-cultured at 26±2°C in dark for 2 d. The maximum transformation efficiency (12%) was achieved with infection of callus explants for 20 min along with use of 150 μm acetosyringone. The maximum plant regeneration was observed on MS medium supplemented with 3 mg/l BAP, 1 mg/l Kinetin and 0.5 mg/l NAA. The maximum root induction was observed on MS medium along with 10 g/l glucose and 20 g/l sucrose. The integration of the transgene in T1 transgenic plants was confirmed by polymerase chain reaction and Southern blot analyses. The copy number of transgenes has been found to vary from 1 to 2 in transgenic plants. By using this improved method we have successfully raised transgenic rice plants within 3 mo from seed inoculation to plant regeneration.     

Shoot regeneration of dwarf dogwood (Cornus canadensis L.) and morphological characterization of the regenerated plants

Dwarf dogwoods (or the bunchberries) are the only suffrutex in Cornaceae. They are attractive ground cover ornamentals with clusters of small flowers surrounded by petaloid bracts. Little has been reported on plant regeneration of dogwoods. As a step toward unraveling the molecular basis of inflorescence evolution in Cornus, we report an efficient regeneration system for a dwarf dogwood species C. canadensis through organogenesis from rejuvenated leaves, and characterize the development of the plantlets. We used the nodal stem segments of vegetative branches as explants. Micropropogated shoots were quickly induced from axillary buds of nodes on an induction medium consisting of basal MS medium supplemented with 4.44 μM BAP and 0.54 μM NAA. The new leaves of adventitious shoots were used as explants to induce calli on the same induction medium. Nearly 65% of leaf explants produced calli, 80% of which formed adventitious buds. Gibberellic acid (1.45 μM) added to the same induction medium efficiently promoted quick elongation of most adventitious buds, and 0.49 μM IBA added to the basal MS medium promoted root formation from nearly 50% of the elongated shoots. The growth of plantlets in pot soil was characterized by the development of functional woody rhizomes, which continuously developed new aboveground vegetative branches, but not flowering branches, within the past 12 months. Potential reasons causing the delay of flowering of the regenerated plants are discussed. The establishment of this regeneration system facilitates developing a genetic transformation system to test candidate genes involved in the developmental divergence of inflorescences in Cornus. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Micropropagation of selected somaclones ofBegonia andSaintpaulia

Begonia x elatior plantlets which regenerated from leaf disk callus showed variations in plant morphology, number of flowers per plant, and flower size. Variations in flowering period, number of flowers per plant, and flower morphology were observed in Saintpaulia ionantha L. plants directly regenerated from leaf disk explants. The cytokinins, benzylaminopurine and zeatin, tested in the culture medium did not affect the basic plant characteristics including flower colour which remained stable in both species. Micropropagation of selected somaclones having the desirable trait of high number of flowers per plant was stable in the MV2 and MV3 generations.     

THE INDUCTION OF FLOWERING IN NICOTIANA. II. PHOTOPERIODIC ALTERATION OF THE CHLOROGENIC ACID CONCENTRATION

Chlorogenic acid and its 2 isomers, believed to be the 4– and 5–0-caffeoylquinic acids, have been extracted from leaves of photoperiodic species of Nicotiana, separated by column chromatography (gradient elution) and measured quantitatively during the course of photoperiodic induction. In both N. tabacum cultivar ‘Maryland Mammoth’ (a short-day plant) and N. sylveslris (a long-day plant), the change of the meristem from the vegetative to the flowering shape is preceded by a rise in the chlorogenic acid content of the leaves. During the differentiation of the flower primordia in the shoot apex, there is a drop in the concentration of the chlorogenic acid in the leaves, which is especially marked in the short-day species. The levels of the 2 isomers decrease also at that time. There is no change in the ratios of the 3 caffeoylquinic acids during photoperiodic induction. In both the long- and the short-day species, the 3-isomer (chlorogenic acid) is present at the highest concentration. In the short-day species, there is more of the 4-isomer than of the 5-isomer, whereas in the long-day species, the level of the 5-isomer equals or surpasses that of the 4–0-caffeoylquinic acid.     

Physiology of Rice Tungro Virus Disease: Involvement of Abscisic Acid-Like Substance in Susceptible Host-Virus Interactions

Stunting was severe in susceptible rice (Oryza saliva L.) cultivar ‘Taichung Native 1’ infected with tungro virus (RTV) compared to less-susceptible cultivar ‘IR 20’. The senescence of detached leaves of RTV-infected susceptible cultivar incubated in water in dark was accelerated compared to the healthy leaves as measured by the loss of total chlorophyll content. The transpiration rate of RTV-infected leaves of the susceptible cultivar was much lower than the healthy and RTV-infected leaves of the less-susceptible cultivar. Partially purified extracts obtained from RTV-infected leaves effectively inhibited GA-induced α-amylase synthesis in barley endosperms, and rice seedling growth, and they accelerated senescence of detached rice leaves. In all the three bibassays the ABA-like activity was significantly greater in the extracts from the RTV-infected susceptible cultivar than in extracts from the less-susceptible cultivar.     

Silicon application promotes rice growth and negatively affects development of Spodoptera frugiperda (J. E. Smith)

Silicon (Si) has been reported to enhance plant resistance against biotic and abiotic stressors and also benefit plant growth. These effects are more pronounced in grass species, especially with soil‐applied Si. This study investigated the effects of Si application on rice resistance to Spodoptera frugiperda development and plant vegetative growth. Effects of Si on rice were assessed via soil and foliar applications and compared with untreated plants (control). Si was soil‐ and foliar‐applied as 1% silicic acid solution at a dosage equivalent to 1.4 t Si per ha. After application, leaf material was collected from Si‐treated and untreated plants and placed in Petri dishes with individual S. frugiperda neonate larvae, where development was followed to adult emergence and biological parameters recorded. Vegetative growth parameters recorded in rice plants were the height, chlorophyll content, fresh and dry weights of shoots, and shoot Si content. No effects of Si application were observed on the durations of larval and pupal stages, larval and pupal survival, and sex ratio of S. frugiperda. Insects fed leaves from Si‐treated plants exhibited lower leaf consumption, larval and pupal weights, longevity of males and females, number of eggs, and egg viability. The negative effects were correlated with higher rice Si content. Si application to rice increased plant height, chlorophyll content and dry weight. Our study demonstrates that foliar‐applied Si is as efficient as soil‐applied Si in negatively affecting S. frugiperda development and providing beneficial effects on rice plant growth.     

Micropropagation of <Emphasis Type="Italic">Theobroma cacao</Emphasis> L. using somatic embryo-derived plants

Summary A micropropagation protocol was developed using cacao somatic embryo-derived plant as a source for nodal and apical stem explants, and apical microcuttings. Microcuttings were efficiently rooted and developed into plantlets. Axillary meristems within the remaining decapitated plantlets subsequently developed and were used for production of additional microcuttings, with an average 2.4 growing shoots per decapitated stem. The remaining plantelts were maintained as microcutting stock plants. When nodal stem explants were cultured on thidiazuron medium, axillary buds proliferated and developed into shoots, which were excised and rooted. However, the efficiency of this method is lower than rooting of apical microcuttings harvested directly from stock plants. During root induction, short treatment with indole-3-butyric acid (IBA) increased the total percentage of rooted microcuttings up to 89%. Longer exposures to IBA increased the average number of roots per microcutting (from 1.7 to 5.2). Plant acclimatization after rooting was achieved with an average success of 87%. During several months of growth in the greenhouse, the micropropagated plants developed functional taproots. Currently, cocoa plants produced by this micropropagation method have been successfully acclimated to field conditions in Ivory Coast, Ghana, and Saint Lucia.     

Growth Enhancement and Developmental Modifications of in Vitro Grown Potato (Solanum tuberosum spp. tuberosum) as Affected by a Nonfluorescent Pseudomonas sp

A plant growth-promoting rhizobacterium, designated Ps JN and isolated from onion roots, was identified as a nonfluorescent Pseudomonas sp. The percentage of similarity of Ps JN to P. gladioli (NCPPB 1891), P. cichorii (NCPPB 943), and P. viridiflava (NCPPB 635), as determined from 135 biochemical and physiological tests was 77, 70, and 66%, respectively. Ps JN persisted through successive generations of in vitro cultured potato plantlets, both as endophytic and epiphytic populations. In vitro inoculated potato (Solanum tuberosum) nodal explants produced plantlets with significant increases in root number (24-196%), root dry weight (44-201%), haulm dry weight (14-151%), and stem length (26-28%) as compared with noninoculated control plants. Bacterization also enhanced leaf hair formation (55-110%), secondary root branching, and total plant lignin content (43%). Other root colonizing bacteria or heat-killed cells of Ps JN had no significant effect on plant growth. Detached leaves from in vitro grown control plants, when exposed to 19°C and 50% relative humidity, lost 55% of their moisture content in 2.5 hours. Moisture loss by leaves of in vitro grown, bacterized plants, as well as greenhouse-acclimated, bacterized plants, and control plants, was less than 20%. Changes in stomatal closure appear to account for this difference.     

Embryogenesis and plant regeneration from tissue culture of Canada wildrye,Elymus canadensis L.

Somatic embryogenesis and plant regeneration of Canada wildrye (Elymus canadensis L.) from tissue culture was investigated by culturing immature embryos and inflorescences on MS medium containing 2 mg/l 2,4-D. The optimum size of explants for maximum embryogenic callus formation was 1.0 to 1.5 mm for embryos and 4 to 6 cm for inflorescences. Plant regeneration from the subcultured embryogenic callus was attempted monthly using hormone-free MS medium or MS medium with 0.5 mg/1 2,4-D and 0.3 mg/l GA3. Three hundred and fifty seven plantlets were regenerated from the callus cultures of both explant sources during a six month period. Ten chlorophyll deficient plants accounting for 2.8% of the total regenerants were observed. One plant with white striped leaves survived and was found to be an octoploid.Abbreviations GA3 gibberellic acid - MS Murashige and Skoog (1962) - 2,4-D 2,4-dichlorophenoxyacetic acid     

A high proportion of blue light increases the photosynthesis capacity and leaf formation rate of Rosa × hybrida but does not affect time to flower opening

Alterations in light quality affect plant morphogenesis and photosynthetic responses but the effects vary significantly between species. Roses exhibit an irradiance‐dependent flowering control but knowledge on light quality responses is scarce. In this study we analyzed, the responses in morphology, photosynthesis and flowering of Rosa × hybrida to different blue (B) light proportions provided by light‐emitting diodes (LED, high B 20%) and high pressure sodium (HPS, low B 5%) lamps. There was a strong morphological and growth effect of the light sources but no significant difference in total dry matter production and flowering. HPS‐grown plants had significantly higher leaf area and plant height, yet a higher dry weight proportion was allocated to leaves than stems under LED. LED plants showed 20% higher photosynthetic capacity (A_max) and higher levels of soluble carbohydrates. The increase in A_max correlated with an increase in leaf mass per unit leaf area, higher stomata conductance and CO₂ exchange, total chlorophyll (Chl) content per area and Chl a/b ratio. LED‐grown leaves also displayed a more sun‐type leaf anatomy with more and longer palisade cells and a higher stomata frequency. Although floral initiation occurred at a higher leaf number in LED, the time to open flowers was the same under both light conditions. Thereby the study shows that a higher portion of B light is efficient in increasing photosynthesis performance per unit leaf area, enhancing growth and morphological changes in roses but does not affect the total Dry Matter (DM) production or time to open flower.