Hypocotyl

Callus induction from petiole explants has been achieved in Alocasia micholitziana `Green Velvet'. The highest percentage (71%) of explants inducing callus was obtained on MS medium supplemented with 0.5 μM 2,4-D and 0.5 μM kinetin in the dark after 4 months of culture. Shoots were regenerated at the highest frequency of 33.3% under light condition when 0.5 μM BA was added to MS medium with the average of 7.8±2.3 shoots per callus explant. The callus-derived shoots rooted on hormone free MS medium and within 4 weeks the plantlets were ready for acclimatization. The regenerated plants appeared morphologically similar to mother plants.

Background: Lepidium meyenii (maca) is a plant that grows exclusively in the Peruvian Central Andes, where ultraviolet radiation (UVR) is predominant. Objective: Determine if two extracts of maca can provide dermal protection against UVR. Methods: We have administered two maca extracts (0.13 mg/ml), one obtained after boiling and the other without boiling, on the dorsal surface of male Holtzman rats exposed to UVC radiation once a week during 3 consecutive weeks. A dose-response effect of an aqueous extract of maca after a boiling process under exposure of rats to UVA, UVB, or UVC was also studied. A commercial sunscreen was used as a positive control. Results: UVR caused significant increase in skin epidermal thickness. The epidermal height in animals treated with maca was similar to those who did not receive UVR. The aqueous extract of maca after a boiling process had better effect than maca extract without a boiling process. A dose-response effect was observed with increasing doses of aqueous extract of maca after a boiling process. Maca extract had benzyl glucosinolates and polyphenols. Conclusions: Maca extracts protect the skin of rats against UV irradiations and can be suggested as an alternative means of solar protection.

Robinia pseudo-acacia L. (black locust) is a nonindigenous species currently invading the central part of Japanese grasslands. Several allelochemicals were identified and characterized from the leaf tissue. The growth of both radicle and hypocotyl in the tested species (barnyard grass, white clover, lettuce, and Chinese cabbage) was reduced when grown in soil mixed with the leaves of R. pseudo-acacia at various concentrations. Aqueous leaf extracts, when bioassayed, exhibited a significant suppression of radicle growth. Chromatographic separation of an ethanolic extract of R. pseudo-acacia leaves resulted in isolation of three compounds, identified as robinetin (1), myricetin (2), and quercetin (3) by nuclear magnetic resonance and mass spectroscopy. All inhibited root and shoot growth of lettuce. Robinetin, found in a large amount, caused 50% suppression of the root and shoot growth of lettuce at 100 ppm. The presence of these bioactive substances in leaf tissue suggests a potential role for flavonoids in R. pseudo-acacia invasion in introduced habitats.

Broccoli inflorescences have been recognized as components of healthy diets on the basis of their high content of fiber, vitamin C, carotenoids, and glucosinolates/isothiocyanates. Broccoli sprouts have been recently shown to have high levels of glucoraphanin (4-methylsulfinylbutyl glucosinolate), the precursor of the chemoprotective isothiocyanate, sulforaphane. This study evaluated the effects of temperature and developmental stage on the glucosinolate content of broccoli sprouts. Seedlings cultivated using a 30/15°C (day/night) temperature regime had significantly higher glucosinolate levels (measured at six consecutive days postemergence) than did sprouts cultivated at lower temperatures (22/15 and 18/12°C; p < 0.001). Both higher (33.1°C) and lower (11.3°C) constant temperatures induced higher glucosinolate levels in sprouts grown to a uniform size. Glucosinolate levels were highest in cotyledons and lowest in roots of sprouts dissected both early and late in the 11 day developmental span investigated. Nongerminated seeds have the highest glucosinolate levels and concordantly greater induction of mammalian phase 2 detoxication enzymes. Levels decline as sprouts germinate and develop, with consistently higher glucosinolate content in younger developmental stages, independent of the temperature regime. Temperature stress or its associated developmental anomalies induce higher glucosinolate levels, specific elevations in glucoraphanin content, and parallel induction of phase 2 chemoprotective enzymes.

The effect of spaceflight on starch development in soybean (Glycine maxL., BRIC-03) and potato (Solanum tuberosum, Astroculture-05) was compared with ground controls by biophysical and biochemical measurements. Starch grains from plants from both flights were on average 20-50% smaller in diameter than ground controls. The ratio A×/Ap (A×difference of magnetic susceptibilities, A p -difference of densities between starch and water) of starch grains was ca. 15% and 4% higher for space-grown soybean cotyledons and potato tubers, respectively, than in corresponding ground controls. Since the densities of particles were similar for all samples (1.36 to 1.38 g/cm3), the observed difference in A~Ap was due to different magnetic susceptibilities and indicates modified composition of starch grains. In starch preparations from soybean cotyledons (BRIC-03) subjected to controlled enzymatic degradation with c~-amylase for 24 hours, 77±6% of the starch from the flight cotyledons was degraded compared to 58+ 12% in ground controls. The amylose content in starch was also higher in space-grown tissues. The good correlation between the amylose content and A~A p suggests, that the magnetic susceptibility of starch grains is related to their amylose content. Since the seedlings from the BRIC-03 experiment showed elevated post-flight ethylene levels, material from another flight experiment (GENEX) which had normal levels of ethylene was examined and showed no difference to ground controls in size distribution, density, A×/Ao and amylose content. Therefore the role of ethylene appears to be more important for changes in starch metabolism than microgravity.

A cost effective procedure was devised for efficient plant propagation of Jatropha curcus L. Direct and indirect in vitro
regeneration was witnessed by addition of plant growth regulator (PGR) combinations. Callus cultures were initiated
from CL, CNR, HC and RT explants on MS basal medium added with 2, 4-D, BAP and NAA in individual and
combined applications. Excellent growth of callus was obtained in all explants with supplementation of BAP (2.0
mg/L) along with NAA (2.0 mg/L), 2, 4-D (0.5 mg/L) being operative for callus induction in only HC explants. These
calli were tested for regeneration and shoot and root organogenesis was successfully induced respectively. Hypocotyl
callus generated 4.0 shoots under the influence of 0.5 mg/L IBA and 1.5 mg/L BAP while CL explant generated 3.0
shoots per callus with slight change in concentration of BAP in same PGRs mixture (0.5 mg/L IBA and 1.0 mg/L
BAP). Along callus mediated organogenic route of regeneration, direct organogenesis was achieved as well. In direct
shoot regeneration ten shoots were formed on CNR with 1.5 mg/L BAP and 0.5 mg/L Kin. Incorporation of 0.25 mg/L
IAA to the medium containing1.5 mg/L BAP + 0.5 mg/L Kin enhanced the regeneration percentage of shoots (11 per
CNR explants). Root initiation in micro-shoots was attained on MS medium (half-strength) added with 0.1 mg/L IBA.
The well-grown plantlets were shifted to peat moss substratum and maintained in field conditions after acclimatization.
This regeneration protocol may also support genetic improvement efforts in J. curcas.

During deetiolation of Arabidopsis seedlings, light promotes the expansion of cotyledons but inhibits the elongation of hypocotyls. The mechanism of this differential regulation of cell enlargement is unclear. Our organ-specific transcriptomic analysis identified 32 Small Auxin Up RNA (SAUR) genes whose transcripts were light-induced in cotyledons and/or repressed in hypocotyls. We therefore named these SAURs as lirSAURs Both overexpression and mutation analyses demonstrated that lirSAURs could promote cotyledon expansion and opening and enhance hypocotyl elongation, possibly by inhibiting phosphatase activity of D-clade type 2C protein phosphatases (PP2C-Ds). Light reduced auxin levels to down-regulate the expression of lirSAURs in hypocotyls. Further, phytochrome-interacting factors (PIFs) were shown to directly bind the genes encoding these SAURs and differentially regulate their expression in cotyledons and hypocotyls. Together, our study demonstrates that light mediates auxin l...

Shoot tips excised from glasshouse-raised seedlings and field-grown plants of cashew (Anacardium occidentale L.) were micrografted by a modified side-grafting procedure using in vitro-raised seedling rootstocks. Factors affecting graft-take were optimised. The micro-scion take obtained was in the range 60-80%. The best growth of micro-scions was obtained with hypocotyls rather than with epicotyls as stock tissue. Rooting of micrografted shoots of mature tree origin was poor (13.3%) because the shoots were only partially rejuvenated.

Seedlings of Cucurbitaceae plants form a protuberance, termed peg, on the transition zone between hypocotyl and root. Our spaceflight experiment verified that the lateral positioning of a peg in cucumber seedlings is modified by gravity. It has been suggested that auxin plays an important role in the gravity controlled positioning of a peg on the ground. Furthermore, cucumber seedlings grown in microgravity developed a number of the lateral roots that grew towards the water containing substrate in the culture vessel, whereas on the ground they oriented perpendicular to the primary root growing down. The response of the lateral roots in microgravity was successfully mimicked by clinorotation of cucumber seedlings on the three dimensional clinostat. However, this bending response of the lateral roots was observed only in an aeroponic culture of the seedlings but not in solid medium. We considered the response of the lateral roots in microgravity and on clinostat as positive hydrotropism that could easily be interfered by gravitropism on the ground. This system with cucumber seedlings is thus a useful model of spaceflight experiment for the study of the gravimorphogenesis, root hydrotropism and their interaction.

Synthetic seeds of cauliflower cv. Chillout were developed by encapsulating mature somatic embryos in neutral gel media. Somatic embryos were obtained by optimizing callus and cell suspension cultures of cauliflower. Friable, yellowish embryogenic calli were obtained on MS supplemented with 2 mg/l 2,4-D and 0.5 mg/l BAP using hypocotyl as explants, while calli were regenerated in media consisting of 5 mg/l BAP, 2 mg/l Kn and 6 mg/l GA3. Somatic embryogenesis was induced in cell suspension culture where auxins were removed in successive steps triggering conversion of globular cells into the heart, torpedo stage (71%) and finally into cotyledonary/somatic embryos (28%). The mature somatic embryos were encapsulated by mixing mature cell suspension with sodium alginate and calcium chloride mixture (1 : 4). Developed synthetic seeds germinated into complete plantlets when placed in neutral gel media. Germination efficiency of synthetic seeds decreased to about 50 per cent after 12 weeks of storage at 4ºC followed by a rapid decrease to zero per cent after 16 weeks. It was also observed that cauliflower plantlets from synthetic seeds survived successfully when transferred to soil demonstrating that cauliflower synthetic seeds is a promising step towards their in vivo direct use.

Knowledge of processes involved in adventitious rooting is important to improve both fundamental understanding of plant physiology and the propagation of numerous plants. Hybrid aspen (Populus tremula 3 tre-mulo€ ıdes) plants overexpressing a key gibberellin (GA) biosynthesis gene (AtGA20ox1) grow rapidly but have poor rooting efficiency, which restricts their clonal propagation. Therefore, we investigated the molecular basis of adventitious rooting in Populus and the model plant Arabidopsis. The production of adventitious roots (ARs) in tree cuttings is initiated from the basal stem region, and involves the interplay of several endogenous and exogenous factors. The roles of several hormones in this process have been characterized, but the effects of GAs have not been fully investigated. Here, we show that a GA treatment negatively affects the numbers of ARs produced by wild-type hybrid aspen cuttings. Furthermore, both hybrid aspen plants and intact Arabidopsis seedlings overexpressing AtGA20ox1, PttGID1.1 or PttGID1.3 genes (with a 35S promoter) produce few ARs, although ARs develop from the basal stem region of hybrid aspen and the hypocotyl of Arabidopsis. In Arabidopsis, auxin and strigolactones are known to affect AR formation. Our data show that the inhibitory effect of GA treatment on adventitious rooting is not mediated by perturbation of the auxin signalling pathway, or of the strigolactone biosynthetic and signalling pathways. Instead, GAs appear to act by perturbing polar auxin transport, in particular auxin efflux in hybrid aspen, and both efflux and influx in Arabidopsis.

A protocol for induction and establishment of Agrobacterium rhizogenes-mediated hairy root cultures of Picrorhiza kurroa was developed through optimization of the explant type and the most suitable bacterial strain. The infection of leaf explants with the LBA9402 strain resulted in the emergence of hairy roots at 66.7% relative transformation frequency. Nine independent, opine and TL-positive hairy root clones were studied for their growth and specific glycoside (i.e., kutkoside and picroside I) productivities at different growth phases. Biosynthetic potentials for the commercially desirable active constituents have been expressed by all the tested hairy root clones, although distinct inter-clonal variations could be noted in terms of their quantity. The yield potentials of the 14-P clone, both in terms of biomass as well as individual glycoside contents (i.e., kutkoside and picroside I), superseded that of all other hairy root clones along with the non-transformed, in vitro-grown control roots of P. kurroa. The present communication reports the first successful establishment, maintenance, growth and selection of superior hairy root clone of Picrorhiza kurroa with desired phyto-molecule production potential, which can serve as an effective substitute to its roots and thereby prevent the indiscriminate up-rooting and exploitation of this commercially important, endangered medicinal plant species.

Plants have evolved shoot elongation mechanisms to escape from diverse environmental stresses such as flooding and vegetative shade. The apparent similarity in growth responses suggests possible convergence of the signalling pathways. Shoot elongation is mediated by passive ethylene accumulating to high concentrations in flooded plant organs and by changes in light quality and quantity under vegetation shade. Here we study hypocotyl elongation as a proxy for shoot elongation and delineated Arabidopsis hypocotyl length kinetics in response to ethylene and shade. Based on these kinetics, we further investigated ethylene and shade-induced genome-wide gene expression changes in hypocotyls and cotyledons separately. Both treatments induced a more extensive transcriptome reconfiguration in the hypocotyls compared to the cotyledons. Bioinformatics analyses suggested contrasting regulation of growth promotion- and photosynthesis-related genes. These analyses also suggested an induction of auxin, brassinosteroid and gibberellin signatures and the involvement of several candidate regulators in the elongating hypocotyls. Pharmacological and mutant analyses confirmed the functional involvement of several of these candidate genes and physiological control points in regulating stress-escape responses to different environmental stimuli. We discuss how these signaling networks might be integrated and conclude that plants, when facing different stresses, utilise a conserved set of transcriptionally regulated genes to modulate and fine tune growth.

Lepidium meyenii) es un hipocotilo peruano que crece exclusivamente entre los 3700 y 4500 msnm en los andes peruanos. Tradicionalmente se le atribuyen propiedades nutricionales, energizantes, fertilizantes, entre otras. Con la finalidad de evaluar científicamente la capacidad nutricional de la Maca, realizamos un estudio controlado en dos generaciones de ratones Swiss (padres y crías). Los padres fueron aleatoriamente asignados a uno de tres esquemas alimenticios. El alimento de cada grupo fue preparado en base a polvo de un alimento balanceado comercial (ABC), al cual se le reemplazó un 30 % (peso/peso) por Maca Cruda o Cocida según el grupo correspondiente o ABC puro en el grupo control. Así los grupos fueron: 1) Grupo Maca Cruda; 2) Grupo Maca Cocida; y 3) Grupo ABC, grupo control. Los resultados mostraron curvas de crecimiento similares y adecuadas para los tres grupos. Ninguno de los grupos mostró signos de desnutrición ni sobrepeso. Sin embargo, el grupo Maca Cocida mostró mejores curvas de crecimiento incluso que el grupo ABC, mejor observable en la segunda generación de animales, con diferencias estadísticamente significativas. El grupo control, tuvo un crecimiento superior al grupo Maca Cruda. Los ratones del grupo Maca Cocida tuvieron valores de proteínas totales y albúmina séricas superiores al de los grupos Maca Cruda y grupo control. Este estudio demuestra en un modelo científico la capacidad nutricional de la Maca.

The scientific literature describing the effects of weak magnetic fields on living systems contains a plethora of contradictory reports, few successful independent replication studies and a dearth of plausible biophysical interaction mechanisms. Most such investigations have been unsystematic, devoid of testable theoretical predictions and, ultimately, unconvincing. A recent study, of magnetic responses in the model plant Arabidopsis thaliana, however, stands out; it has a clear hypothesis-that seedling growth is magnetically sensitive as a result of photoinduced radical-pair reactions in cryptochrome photoreceptors-tested by measuring several cryptochrome-dependent responses, all of which proved to be enhanced in a magnetic field of intensity 500 mT. The potential importance of this study in the debate on putative effects of extremely low-frequency electromagnetic fields on human health prompted us to subject it to the 'gold standard' of independent replication. With experimental conditions chosen to match those of the original study, we have measured hypocotyl lengths and anthocyanin accumulation for Arabidopsis seedlings grown in a 500 mT magnetic field, with simultaneous control experiments at 50 mT. Additionally, we have determined hypocotyl lengths of plants grown in 50 mT, 1 mT and approximately 100 mT magnetic fields (with zero-field controls), measured gene (CHS, HY5 and GST ) expression levels, investigated blue-light intensity effects and explored the influence of sucrose in the growth medium. In no case were consistent, statistically significant magnetic field responses detected.

Caffeic acid (CA) is one of the most common cinnamic acids ubiquitously present in plants and implicated in a variety of interactions including allelopathy among plants and microbes. This study investigated the possible interference of CA with root growth and the process of rhizogenesis in hypocotyl cuttings of mung bean (Phaseolus aureus=Vigna radiata). Results indicated that CA (0–1000 μM) significantly suppressed root growth of mung bean, and impaired adventitious root formation and root length in the mung bean hypocotyl cuttings. Further investigations into the role of CA in hampering root formation indicated its interference with the biochemical processes involved in rooting process at the three stages – root initiation (third day; RI), root expression (fifth day; RE), and post-expression (seventh day; PE) – of rhizogenesis. CA caused significant changes in the activities of proteases, peroxidases (PODs), and polyphenol oxidases (PPOs) during root development and decreased the content of total endogenous phenolics (TP) in the hypocotyl cuttings. The enhanced activity of PODs and PPOs, though, relates to lignification and/or phenolic metabolism during rhizogenesis; yet their protective role to CA-induced stress, especially during the PE phase, is not ruled out. At 1000 μM CA, where rooting was significantly affected, TP content was very high during the RI phase, thus indicating its non-utilization. The study concludes that CA interferes with the rooting potential of mung bean hypocotyl cuttings by altering the activities of PODs and PPOs and the endogenous TP content that play a key role in rhizogenesis.

Efficient plant regeneration protocol through callus for Saussurea obvallata (DC.) Edgew. (Asteraceae): effect of explant type, age and plant growth regulators Abstract A callus induction and in vitro plantlet regeneration system for the endangered state flower of Uttaranchal (Saussurea obvallata) was optimized by studying the influence of explant type (root, hypocotyl, cotyledon and leaf), age and different concentrations of plant growth regulators. Explants from 10 to 15-day-old seedlings showed maximum callus induction. Callus formation and shoot differentiation was initiated on Murashige-Skoog (MS) medium containing 6-benzyladenine (BA) and α-naphthalene acetic acid (NAA) in all explant types. The best results were obtained using leaf explants: 100% callusing was achieved in MS medium supplemented with 2.5 µM BA and 1.0 µM NAA, and 100% differentiation along with a multiplication rate of 12 shoots per explant with a combination of 5.0 µM BA and 1.0 µM NAA. However, the results reflected the existence of high inter-explant variability in response to growth regulators. In vitro rooting of shoots was achieved at an efficiency of 100% in one-half strength MS medium supplemented with 2.5 µM indole-3butyric acid. Application of this protocol has potential for mass multiplication of the target species in a limited time period.

PHYTOCHROME-INTERACTING FACTOR5 (PIF5), a basic helix-loop-helix transcription factor, interacts specifically with the photoactivated form of phytochrome B (phyB). Here, we report that dark-grown Arabidopsis thaliana seedlings overexpressing PIF5 (PIF5-OX) exhibit exaggerated apical hooks and short hypocotyls, reminiscent of the triple response induced by elevated ethylene levels, whereas pif5 mutants fail to maintain tight hooks like those of wild-type seedlings. Silver ions, an ethylene receptor blocker, rescued the triple-response phenotype, and we show that PIF5-OX seedlings express enhanced levels of key ethylene biosynthesis enzymes and produce elevated ethylene levels. Exposure of PIF5-OX seedlings to prolonged continuous red light (Rc) promotes hypocotyl elongation relative to dark controls, the reciprocal of the Rc-imposed hypocotyl inhibition displayed by wild-type seedlings. In contrast with this PIF5-OX hyposensitivity to Rc, pif5 mutant seedlings are hypersensitive rela...

Five different varieties of Brassica napus (Cyclone, Dunkled, Oscar, Rainbow and KS75) were tested for their regeneration response. Cyclone showed a very high frequency of regeneration (92%). The use of silver nitrate was a pre-requisite for efficient shoot regeneration. Hypocotyls were selected as the starting material for transformation experiments on the basis of high transient GUS expression. Explants were co-cultivated with Agrobacterium strain EHA101 harboring a binary vector pIG121Hm containing neomycin phosphotransferase II (NPTII) gene, conferring resistance to kanamycin, hygromycin phosphotransferase (HPT) gene, conferring resistance to hygromycin as selectable markers and β-glucuronidase (GUS) gene as a reporter. Acetosyringone promoted the transformation but was not an absolute requirement. A pre-selection period of 7 days after co-cultivation was essential for successful transformation. Kanamycin was efficient selective agent for selection and maximum transformation efficiency was 24%. GUS activity was evident in leaf tissues. All the transgenic plants have an expected band of 0.43 kb fragment by PCR analysis confirming the presence of foreign DNA into plant genome.

Life occurs in an ever-changing environment. Some of the most striking and predictable changes are the daily rhythms of light and temperature. To cope with these rhythmic changes, plants use an endogenous circadian clock to adjust their growth and physiology to anticipate daily environmental changes. Most studies of circadian functions in plants have been performed under continuous conditions. However, in the natural environment, diurnal outputs result from complex interactions of endogenous circadian rhythms and external cues. Accumulated studies using the hypocotyl as a model for plant growth have shown that both light signalling and circadian clock mutants have growth defects, suggesting strong interactions between hypocotyl elongation, light signalling and the circadian clock. Here, we review evidence suggesting that light, plant hormones and the circadian clock all interact to control diurnal patterns of plant growth.

The shade avoidance syndrome (SAS) refers to a set of plant responses initiated after perception by the phytochromes of light with a reduced red to far-red ratio, indicative of vegetation proximity or shade. These responses, including elongation growth, anticipate eventual shading from potential competitor vegetation by overgrowing neighboring plants or flowering to ensure production of viable seeds for the next generation. In Arabidopsis thaliana seedlings, the SAS includes dramatic changes in gene expression, such as induction of PHYTOCHROME RAPIDLY REGULATED 1 (PAR1), encoding an atypical basic helix-loop-helix (bHLH) protein that acts as a transcriptional co-factor to repress hypocotyl elongation. Indeed, PAR1 has been proposed to act fundamentally as a dominant negative antagonist of conventional bHLH transcription factors by forming heterodimers with them to prevent their binding to DNA or other transcription factors. Here we report the identification of PAR1-interacting factors, including the brassinosteroid signaling components BR-ENHANCED EXPRESSION (BEE) and BES1-INTERACTING MYC-LIKE (BIM), and characterize their role as networked positive regulators of SAS hypocotyl responses. We provide genetic evidence that these bHLH transcriptional regulators not only control plant growth and development under shade and non-shade conditions, but are also redundant in the control of plant viability. Our results suggest that SAS responses are initiated as a consequence of a new balance of transcriptional regulators within the pre-existing bHLH network triggered by plant proximity, eventually causing hypocotyls to elongate.

We analyzed the natural genetic variation between Landsburg erecta (Ler) and Cape Verde Islands (Cvi) accessions by studying 105 recombinant inbred lines to search for players in the regulation of sensitivity to light signals perceived by phytochromes in etiolated seedlings of Arabidopsis. In seedlings grown under hourly pulses of far-red (FR) light, we identified three quantitative trait loci (QTLs; VLF3, VLF4, and VLF5) for hypocotyl growth inhibition and three different QTLs (VLF6, VLF7, and VLF1) for cotyledon unfolding. This indicates that different physiological outputs have selective regulation of sensitivity during de-etiolation. Ler alleles, compared with Cvi alleles, of VLF3, VLF4, VLF5, VLF7, and VLF1 enhanced, whereas the Ler allele of VLF6 reduced, the response to pulses of FR. We confirmed and narrowed down the position of some QTLs by using near-isogenic lines. VLF6 mapped close to the CRY2 (cryptochrome 2) gene. Transgenic Ler seedlings expressing the Cvi allele of CRY2 showed enhanced cotyledon unfolding under hourly pulses of FR compared with the wild type or transgenics expressing the CRY2-Ler allele. This response required phytochrome A. The cry1 cry2 double mutant lacking both cryptochromes showed reduced cotyledon unfolding under FR pulses. Because the CRY2-Cvi is a gain-of-function allele compared with CRY2-Ler, cryptochrome activity correlates positively with cotyledon unfolding under FR pulses. We conclude that the blue light photoreceptor cryptochrome 2 can modulate seedling photomorphogenesis in the absence of blue light. In addition to the nuclear loci, we identified cytoplasmic effects on seedling de-etiolation. fax 54 -011-4514 -8730.

The effect of smoke saturated-water (SSW) on somatic embryogenesis was studied using geranium hypocotyl culture as a model system. Treatment of explants with 10% SSW or the inclusion of SSW with thidiazuron, a compound which induces somatic embryogenesis, enhanced the embryogenic potential of the geranium hypocotyl culture. Prolonged exposure to SSW was detrimental to embryogenesis. The SSW treatment also accelerated the rate of embryo development suggesting a growth regulatory role of SSW.

Phototropin 1 (phot1) is a photoreceptor for phototropism, chloroplast movement, stomatal opening, leaf expansion, and solar tracking in response to blue light. Following earlier work with PHOT1:: GFP (Sakamoto and Briggs, 2002), we investigated the pattern of cellular and subcellular localization of phot1 in 3-4 d old etiolated seedlings of Arabidopsis thalinana. As expressed from native upstream sequences, the PHOT1::GFP fusion protein is expressed strongly in the abaxial tissues of the cotyledons and in the elongating regions of the hypocotyl. It is moderately expressed in the shoot/root transition zone and in cells near the root apex. A fluorescence signal is undetectable in the root epidermis, root cap, and root apical meristem itself. The plasma membranes of mesophyll cells near the cotyledon margin appear labeled uniformly but cross-walls created by recent cell divisions are more strongly labeled. The pattern of labeling of individual cell types varies with cell type and developmental stage. Blue-light treatment causes PHOT1::GFP, initially relatively evenly distributed at the plasma membrane, to become reorganized into a distinct mosaic with strongly labeled punctate areas and other areas completely devoid of fluorescence-a phenomenon best observed in cortical cells in the hypocotyl elongation region. Concomitant with or following this reorganization, PHOT1::GFP moves into the cytoplasm in all cell types investigated except for guard cells. It disappears from the cytoplasm by an unidentified mechanism after several hours in darkness. Neither its appearance in the cytoplasm nor its eventual disappearance in darkness is prevented by the translation inhibitor cycloheximide, although the latter process is retarded. We hypothesize that blue-light-induced phot1 relocalization modulates blue-light-activated signal transduction.

Plants modify growth in response to the proximity of neighbors. Among these growth adjustments are shade avoidance responses, such as enhanced elongation of stems and petioles, that help plants to reach the light and outgrow their competitors. Neighbor detection occurs through photoreceptor-mediated detection of light spectral changes (i.e. reduced red:far-red ratio [R:FR] and reduced blue light intensity). We recently showed that physiological regulation of these responses occurs through light-mediated degradation of nuclear, growth-inhibiting DELLA proteins, but this appeared to be only part of the full mechanism. Here, we present how two hormones, auxin and ethylene, coregulate DELLAs but regulate shade avoidance responses through DELLA-independent mechanisms in Arabidopsis (Arabidopsis thaliana). Auxin appears to be required for both seedling and mature plant shoot elongation responses to low blue light and low R:FR, respectively. Auxin action is increased upon exposure to low R:FR and low blue light, and auxin inhibition abolishes the elongation responses to these light cues. Ethylene action is increased during the mature plant response to low R:FR, and this growth response is abolished by ethylene insensitivity. However, ethylene is also a direct volatile neighbor detection signal that induces strong elongation in seedlings, possibly in an auxin-dependent manner. We propose that this novel ethylene and auxin control of shade avoidance interacts with DELLA abundance but also controls independent targets to regulate adaptive growth responses to surrounding vegetation.

We have used a well-characterized antibody specific for an epitope consisting of (1→3,6)-β-D-galactosyl residues with terminal glucuronic or 4-O-methylglucuronic acids of a bioactive pectin and immunocytochemistry to investigate its secretion and wall distribution in the hypocotyl and root tissues of flax seedlings. Our results show that this antigenic epitope is associated with flax pectins and is expressed by all the cells of the hypocotyl and root tissues. In the hypocotyl, it is abundant in the primary wall of epidermal cells as well as in the secondary wall of fiber cells, and is relatively less abundant in parenchyma cell walls. In contrast, the epitope is not detected in the middle lamellae and cell junction regions. In the root tip cells, immunogold electron microscopy shows that the cell walls of peripheral, columella, meristematic, cortical, and epidermal cells contain significant amounts of this epitope and that the distribution patterns are distinct. Together, these findings show that the antigenic epitope occurs in discrete domains of the wall implying a strict spatial regulation of the epitope-containing molecules. The results also show that, in root cells, the epitope is present within Golgi cisternae and is predominantly assembled in the trans and the trans-Golgi network compartments.

CONSTITUTIVE PHOTOMORPHOGENIC1 (COP1) is an E3 ubiquitin ligase that represses photomorphogenesis in the dark. Therefore, proteins interacting with COP1 could be important regulators of light-dependent development. Here, we identify CONSTANS-LIKE3 (COL3) as a novel interaction partner of COP1. A green fluorescent protein-COL3 fusion protein colocalizes with COP1 to nuclear speckles when transiently expressed in plant cells. This localization requires the B-box domains in COL3, indicating a novel function of this domain. A loss-of-function col3 mutant has longer hypocotyls in red light and in short days. Unlike constans, the col3 mutant flowers early and shows a reduced number of lateral branches in short days. The mutant also exhibits reduced formation of lateral roots. The col3 mutation partially suppresses the cop1 and deetiolated1 (det1) mutations in the dark, suggesting that COL3 acts downstream of both of these repressors. However, the col3 mutation exerts opposing effects on cop1 and det1 in terms of lateral roots and anthocyanin accumulation, suggesting that COL3 also has activities that are independent of COP1 and DET1. In conclusion, we have identified COL3 as a positive regulator of photomorphogenesis that acts downstream of COP1 but can promote lateral root development independently of COP1 and also function as a daylength-sensitive regulator of shoot branching.

Melatonin (N-acetyl-5-methoxi-tryptamine), a well-known animal hormone synthetised by the pineal gland, plays a key role in the circadian rhythm of vertebrates. An exhaustive bibliographical revision of studies on melatonin in plants published since 1990 points to very few studies (around 20), of which only 8 have a clear plant physiological focus. The data presented in this study demonstrate that melatonin plays a physiological role in plant tissues. Melatonin is seen to be a molecule that promotes vegetative growth in etiolated Lupinus albus L. hypocotyls, in a similar way to IAA. The measurements of melatonin and IAA in lupin hypocotyls by high-performance liquid chromatography with electrochemical detection, and their identification by tandem mass spectrometry, point to a different distribution of these molecules in etiolated hypocotyls.

Fusarium wilt, caused by Fusarium oxysporum (Fo), is one of the most important fungal diseases worldwide. Like other plant pathogens, Fo displays specialized forms in association with its hosts. For example, F. oxysporum f. sp. niveum (Fon) is the damaging pathogen causing Fusarium wilt disease on watermelon, whereas F. oxysporum f. sp. cubense is the pathogen that infects banana. A rapid and reliable pathogen identification or disease diagnosis is essential for the integrated disease management practices in many crops. In this study, two new primer sets, Fon-1/Fon-2 and FnSc-1/FnSc-2, were developed to differentiate Fon and Fo, respectively. The PCR method using the novel primer sets has high sensitivity to detect Fon when the DNA concentration was as low as 0.01 pg or when the conidia number was as few as 5. In comparison with the published primer set, the Fon-1/Fon-2 primer set, derived from the sequence of OP-M12 random primer-amplified fragment, produced a 174 bp DNA fragment, and was more specific to Fon in Taiwan. In addition, with optimized PCR parameters, the molecular method using the Fon-1/Fon-2 primer set could directly detect Fon even when watermelon samples were collected in its early stage of disease development.

Background and AimsPlants growing at high densities perceive a decrease in the red to far-red (R/FR) ratio of incoming light. These changes in light quality trigger a suite of responses collectively known as the shade-avoidance syndrome (SAS) including hypocotyl and stem elongation, inhibition of branching and acceleration of flowering.Methods Quantitative trait loci (QTLs) were mapped for hypocotyl length to end-of-day far-red (EOD), a simulated shade-avoidance response, in recombinant inbred line (RIL) populations of Arabidopsis thaliana seedlings, derived from Landsberg erecta (Ler) and three accessions (Columbia, Col; Nossen, No-0; and Cape Verde Islands, Cvi-0).Key ResultsFive loci were identified as being responsible for the EOD response, with a positive contribution of Ler alleles on the phenotype independently of the RIL population. Quantitative complementation analysis and transgenic lines showed that PHYB is the candidate gene for EODRATIO5 in the Ler × Cvi-0 RIL populatio...

We studied the effect of elevated boron (B) concentrations on the growth and development of Arabidopsis thaliana in vitro with respect to different light conditions. Two basic responses were observed. At high concentrations (above 5 mM) a clear toxicity effect of B on plant growth was apparent. Seedlings were short, stunted and pale. However at concentrations between 1 and 3 mM H 3 BO 3 , hypocotyl elongation was stimulated in all Arabidopsis ecotypes tested relative to plants grown at 0.1 mM H 3 BO 3. The stimulation of hypocotyl elongation by elevated B was proportionally greater with increasing irradiance. We also showed that blue light (BL) and red light (RL) did not alter the sensitivity of Arabidopsis hypocotyls to boron, but, dependent on genotype, BL and RL increased or reduced capacity of boron-induced hypocotyl elongation. Analysis of photomorphogenic mutants indicated the existence of an interaction between boron and light signalling pathways during plant growth and development. This interaction was supported by the observation that the expression of the BOR1 gene in Arabidopsis hypocotyls was stimulated by BL and RL. Our results suggest that in etiolated or light-grown seedlings the stimulation of hypocotyl growth by boron can be mediated by cryptochromes and phytochromes.

The pathogenicity, host range and infection process of three isolates of Colletotrichum capsici , from cowpea (Vigna unguiculata), bean (Phaseolus vulgaris) and betle vine (Piper betle), are described. The location and morphology of infection hyphae of C. capsici within hypocotyls of cowpea were investigated by light, transmission and cryo-scanning electron microscopy. All isolates attacked a similar range of legumes, but

Unlike other eukaryotes, which synthesize polyamines (PA) only from ornithine, plants possess an additional pathway utilizing arginine as a precursor. In this study, we have identified cDNA clones coding for a Glycine max ornithine decarboxylase (ODC, EC 4.1.1.7) and an arginine decarboxylase (ADC, EC 4.1.1.19). Expression analysis using semi-quantitative RT-PCR approach revealed that both genes coding for enzymes involved in putrescine biosynthesis (ODC and ADC) were found in most plant organs examined. Significant expression levels of both genes were detected in root tips and hypocotyls. The spatial distribution of GmODC and GmADC transcripts in primary and lateral roots and hypocotyls revealed that these genes are co-expressed in expanding cells of cortex parenchyma, expanding cells of central cylinder in main roots and in developing tissues and expanding cells of soybean hypocotyls. The data point out a correlation of the expression patterns of GmODC and GmADC gene to certain physiological roles such as organ development and cell expansion.

Although several types of plant cells retain the competence to enter into embryonic development without fertilization, the molecular mechanism(s) underlying ectopic embryogenesis is largely unknown. To gain insight into this mechanism, in a previous study we identified 136 ESTs specifically expressed in microspore embryogenesis of Brassica napus. Here, we describe the characterization of the Arabidopsis EMBRYOMAKER (EMK) gene, which is homologous to one of the identified Brassica ESTs (BnGemb-58) and encodes an AP2 domain transcription factor. The AtEMK was expressed in developing and mature embryos, but its rapid disappearance occurred during germination. After germination, the expression of AtEMK was found in the root apical meristem and the distal parts of cotyledons. Although a mutant lacking AtEMK exhibited no distinctive defects in the embryo, ectopic expression of AtEMK induced embryo-like structures from cotyledons. The embryo-like structures contained high concentration of lipids, expressed several embryo-specific genes, and could convert into independent plants, indicating that the structures are somatic embryos. In vitro culture, AtEMK enhanced the efficiency of somatic embryogenesis. Furthermore, ectopic expression of AtEMK caused the formation of trichomes on cotyledons, dedifferentiated several tissues into calli, and retarded root development, demonstrating that AtEMK is harmful for the normal development of plants after germination. From these results, we conclude that the AtEMK is a key player to maintain embryonic identity, and the rapid disappearance of AtEMK expression during germination is essential for the developmental transition between the embryonic and vegetative phases in plants.

3-Hydroxy-4,5-dimethyl-2(5H)-furanone (sotolone) and 3-amino-4,5-dimethyl-2(5H)-furanone, the postulated precursor of sotolone, were detected in hairy root cultures of Trigonella foenum-graecum (fenugreek) by GC-MS. The hairy root cultures in both conical flasks and airlift with mesh bioreactors were achieved from hypocotyl of seedling by infection with Agrobacterium rhizogenes. In flasks, the mathematical relationship between hairy root growth and conductivity was established and afterward used to evaluate the biomass evolution in bioreactor cultures due to the difficulty of obtaining direct biomass samples from the bioreactor. The GC-MS analyses of ethanolic extracts from hairy roots revealed the presence of two important compounds: sotolone (1.2% of the volatile fraction) and 3-amino-4,5-dimethyl-2(5H)-furanone (17% of the volatile fraction). These results point out that biotechnological production of sotolone in bioreactors is possible. Additionally, these hairy root cultures offer, for the first time, an excellent biological model to study the biosynthetic pathway of sotolone in fenugreek.

The growth direction of the Arabidopsis (Arabidopsis thaliana) etiolated-seedling hypocotyl is a complex trait that is controlled by extrinsic signals such as gravity and touch as well as intrinsic signals such as hormones (brassinosteroid [BR], auxin, cytokinin, ethylene) and nutrient status (glucose [Glc], sucrose). We used a genetic approach to identify the signaling elements and their relationship underlying hypocotyl growth direction. BR randomizes etiolated-seedling growth by inhibiting negative gravitropism of the hypocotyls via modulating auxin homeostasis for which we designate as reset, not to be confused with the gravity set point angle. Cytokinin signaling antagonizes this BR reset of gravity sensing and/or tropism by affecting ethylene biosynthesis/signaling. Glc also antagonizes BR reset but acts independently of cytokinin and ethylene signaling pathways via inhibiting BR-regulated gene expression quantitatively and spatially, by altering protein degradation, and by an...