Morphogenic Responses Research Articles

Low Cd concentration-activated morphogenic defence responses are inhibited by high Cd concentration-induced toxic superoxide generation in barley root tip

Exposure of roots to low Cd concentration induced morphogenic responses including the inhibition of root growth and the radial swelling of root tip. High Cd concentrations within a few minutes caused a robust induction of superoxide generation leading to the cell death and root growth arrest. This toxic superoxide generation blocked the development of low Cd concentration-activated morphogenic responses. While the morphogenic responses of roots to low Cd concentration are induced very rapidly and probably due to the interaction of Cd with the apoplast of root tissue, high Cd concentration-induced superoxide production required the entry of Cd into the symplast. Auxin signaling is involved in the activation of Cd-induced morphogenic defence responses but not in the Cd-induced toxic superoxide generation. These results suggest that oxidative stress is not a primary cause for the Cd-induced morphogenic responses such as growth reduction and radial cell expansion in barley root tips.

Plant adaptation to extreme environments: The example of Cistus salviifolius of an active geothermal alteration field

Cistus salviifolius is able to colonise one of the most extreme active geothermal alteration fields in terms of both soil acidity and hot temperatures. The analyses of morpho-functional and physiological characters, investigated in leaves of plants growing around fumaroles (G leaves) and in leaves developed by the same plants after transfer into growth chamber under controlled conditions (C leaves) evidenced the main adaptive traits developed by this pioneer plant in a stressful environment. These traits involved leaf shape and thickness, mesophyll compactness, stomatal and trichome densities, chloroplast size. Changes of functional and physiological traits concerned dry matter content, peroxide and lipid peroxidation, leaf area, relative water and pigment contents. A higher reducing power and antioxidant enzymatic activity were typical of G leaves. Though the high levels of stress parameters, G leaves showed stress-induced specific morphogenic and physiological responses putatively involved in their surviving in active geothermal habitats.

In vitro morphogenic response of different explants of Gentiana kurroo Royle from Western Himalayas-an endangered medicinal plant.

Micropropagation offers a great potential to produce millions of clonal individuals through tissue culture via induction of morphogenesis. The aim of this work was to obtain an efficient protocol for callus regeneration for Gentiana kurroo Royle. The morphogenic response of different explants (leaves, petioles, roots) varied and responded differently for regeneration according to combinations of growth regulators. The petiole explants were best responding for callus induction and subsequently for indirect and direct regeneration. The callus induction was achieved on MS basal + 1.0mg/l benzyladenine (BA) and 3.00mg/l naphthalene acetic acid (NAA). MS medium supplemented with 0.10mg/l NAA and 1.0mg/l thidiazuron (TDZ) was recorded as the best medium for indirect regeneration. However, for direct regeneration the maximum number of shoot emergence was observed on MS basal fortified with 0.10mg/l NAA + 0.75mg/l TDZ. Half strength MS basal supplemented with indole-3-butyric acid (IBA) 1.00mg/l gave best response for root induction. Subsequently, the plantlets were transferred and 100% survival rate was recorded only on autoclaved cocopeat. No morphological variations were recorded in the callus regenerated plantlets.

L-arginine impact on cherry rootstock rooting and biochemical characteristics in tissue culture

In the present study, the effects of indole-3-butyric acid (IBA) separately and simultaneously with L-arginine on the morphogenic and biochemical responses in the cherry rootstocks CAB-6P (Prunus cerasus L.) and Gisela 6 (Prunus cerasus × Prunus canescens) were investigated. In the CAB-6P rootstock, the best root number and root length results were obtained with 2 mg L–1 IBA plus 0.5 mg L–1 L-arginine and 1 mg L–1 IBA plus 1 mg L–1 L-arginine, respectively. The rooting percentage was highest (100%) with 2 mg L–1 IBA alone or combined with 1 mg L–1 L-arginine. In the Gisela 6 explants, 2 mg L–1 IBA without L-arginine significantly enhanced root number (9.27) and root fresh and dry weight, as well as rooting percentage (100%). Root length was longest (38 mm) in the combination of the lowest IBA (0.5 mg L–1) and highest L-arginine (2 mg L–1) concentrations. Thus, L-arginine promotes the positive effect of IBA on rooting with regard to both root number and root length in both cherry rootstocks. In the CAB-6P explants, L-arginine combined with IBA had an inhibitory effect on leaf chlorophyll content, whereas in the Gisela 6 rootstock it had absolutely no effect. In the CAB-6P rootstock, depleted levels of proline in roots were observed, showing the osmoregulation and osmotic adjustment mechanisms. The carbohydrate concentration in leaves was greatest with 0.5 mg L–1 IBA plus 0.5 mg L–1 L-arginine, and in roots with 2 mg L–1 IBA plus 0.5 mg L–1 L-arginine. In the Gisela 6 explants, IBA (1 and 2 mg L–1) applied both alone and along with L-arginine, as well as the combination of the lowest IBA (0.5 mg L–1) and the highest L-arginine (2 mg L–1) concentrations, considerably increased leaf carbohydrate concentration compared to the control. In roots, carbohydrate concentration was maximum with 2 mg L–1 IBA plus 0.5 mg L–1 L-arginine. The elevated levels of proline in roots with 1 mg L–1 IBA plus 0.5 mg L–1 L-arginine indicate stressful conditions.

A Detailed Comparative Investigation of Morphogenic Responses of Varied Phytohormones inMimosa pudicaandTinospora cordifolia

In this communication, an attempt has been made to study a comparative analysis of morphogenic responses between two species viz. Mimosa pudica (fam. Mimosaceae) and Tinospora cordifolia (Fam. Menispermaceae). Effects of phytohormones viz. auxins (IAA and NAA), cytokinins (BAP and Kinetin) and gibberellins have been recorded in both species. Differentiation of shoots and roots have been encountered while formation of callusing has been marked out. Mimosa pudica appears to be more responsive in comparision to Tinospora cordifolia.

Allelic Interaction between CRELD1 and VEGFA in the Pathogenesis of Cardiac Atrioventricular Septal Defects

Atrioventricular septal defects (AVSD) are highly heritable, clinically significant congenital heart malformations. Genetic and environmental modifiers of risk are thought to work in unknown combinations to cause AVSD. Approximately 5–10% of simplex AVSD cases carry a missense mutation in CRELD1. However, CRELD1 mutations are not fully penetrant and require interactions with other risk factors to result in AVSD. Vascular endothelial growth factor-A (VEGFA) is a well-characterized modulator of heart valve development. A functional VEGFA polymorphism, VEGFA c.–634C, which causes constitutively increased VEGFA expression, has been associated with cardiac septal defects suggesting it may be a genetic risk factor. To determine if there is an allelic association with AVSD we genotyped the VEGFA c.–634 SNP in a simplex AVSD study cohort. Over-representation of the c.–634C allele in the AVSD group suggested that this genotype may increase risk. Correlation of CRELD1 and VEGFA genotypes revealed that potentially pathogenic missense mutations in CRELD1 were always accompanied by the VEGFA c.–634C allele in individuals with AVSD suggesting a potentially pathogenic allelic interaction. We used a Creld1 knockout mouse model to determine the effect of deficiency of Creld1 combined with increased VEGFA on atrioventricular canal development. Morphogenic response to VEGFA was abnormal in Creld1-deficient embryonic hearts, indicating that interaction between CRELD1 and VEGFA has the potential to alter atrioventricular canal morphogenesis. This supports our hypothesis that an additive effect between missense mutations in CRELD1 and a functional SNP in VEGFA contributes to the pathogenesis of AVSD.

Plantlets from encapsulated shoot buds of Catalpa ovata G. Don

Shoot buds isolated from in vitro shoot cultures of <em>Catalpa ovata</em> G. Don were encapsulated using 3% sodium alginate with sucrose (3%) and 50 mM calcium chloride. The morphogenic response of encapsulated buds was affected by such factors, like composition of the media and the presence of growth regulators. The highest frequency of plantlet germination from encapsulated buds (70% within 4 weeks) was obtained on Woody Plant medium (WP) (Lloyd and McCown 1980) containing indole-3-butyric acid (IBA) (1 mg/l). The process was substantially inhibited by cold-storage (4<sup>o</sup>C) of encapsulated buds. In this case, the frequency response ranged from 3% to 22% dependent on storage period (28 or 42 days) and the presence of the paraffin coat covering the alginate capsules. The plantlets developed from both unstored and stored encapsulated buds of <em>C. ovata</em> were transplanted to soil and grew in pots to phenotypically normal plants.

Hematopoietic Stem Cell Cytokines and Fibroblast Growth factor-2 Stimulate Human Endothelial Cell-Pericyte Tube Co-Assembly in 3D Fibrin Matrices under Serum-Free Defined Conditions

We describe a novel 3D fibrin matrix model using recombinant hematopoietic stem cell cytokines under serum-free defined conditions which promotes the assembly of human endothelial cell (EC) tubes with co-associated pericytes. Individual ECs and pericytes are randomly mixed together and EC tubes form that is accompanied by pericyte recruitment to the EC tube abluminal surface over a 3-5 day period. These morphogenic processes are stimulated by a combination of the hematopoietic stem cell cytokines, stem cell factor, interleukin-3, stromal derived factor-1α, and Flt-3 ligand which are added in conjunction with fibroblast growth factor (FGF)-2 into the fibrin matrix. In contrast, this tube morphogenic response does not occur under serum-free defined conditions when VEGF and FGF-2 are added together in the fibrin matrices. We recently demonstrated that VEGF and FGF-2 are able to prime EC tube morphogenic responses (i.e. added overnight prior to the morphogenic assay) to hematopoietic stem cell cytokines in collagen matrices and, interestingly, they also prime EC tube morphogenesis in 3D fibrin matrices. EC-pericyte interactions in 3D fibrin matrices leads to marked vascular basement membrane assembly as demonstrated using immunofluorescence and transmission electron microscopy. Furthermore, we show that hematopoietic stem cell cytokines and pericytes stimulate EC sprouting in fibrin matrices in a manner dependent on the α5β1 integrin. This novel co-culture system, under serum-free defined conditions, allows for a molecular analysis of EC tube assembly, pericyte recruitment and maturation events in a critical ECM environment (i.e. fibrin matrices) that regulates angiogenic events in postnatal life.

Ethylene Modulates the Developmental Plasticity and the Growth Balance Between Shoot and Root Systems in the In Vitro Grown Epiphytic Orchid Catasetum fimbriatum

The epiphytic habitat is potentially one of the most stressful environments for plants, making the effective developmental control in response to external cues critical for epiphyte survival. Because ethylene mediates several abiotic stresses in plants, here, we have examined the ethylene influence in both shoot and root systems of the epiphytic orchid Catasetum fimbriatum. Under controlled conditions, ethylene production was quantified during an entire growth cycle of C. fimbriatum development in vitro, while treatments modulating either ethylene concentration or perception were carried out over the early growth phase of these plants. After treatments, growth measurements and histological features were studied in both shoot and root tissues. Ethylene production showed a decreasing trend over the period of organ elongation; however, it increased considerably when leaves were shed, and a new axillary bud was initiating. The early exposure of young plants to higher concentrations of ethylene triggered morphogenic responses that included root hair formation instead of velamen, and a combination of inhibitory effects (decreases in both stem enlargement and cellular/organ elongation) and inductive effects (increases in leaf and root formation, bud initiation and cellular thickening) on plant growth, which favored biomass allocation to roots. Conversely, inhibition of ethylene perception over the plant growth phase generally resulted in the opposite morphogenic responses. Our data indicate that periodic variations in ethylene concentration and/or sensitivity seem to modulate several developmental features in shoot and root systems of C. fimbriatum which could have adaptive significance during the growing phase of this epiphytic orchid.

Suppression of ethylene levels promotes morphogenesis in pepper (Capsicum annuum L.)

Ethylene and polyamines (PAs) are two phytohormones that play important roles during in vitro morphogenesis of several plant species. The interaction between ethylene and PAs has been of interest because both have S-adenosylmethionine as a precursor. To study the influence of ethylene and PAs on in vitro morphogenesis of an ornamental pepper, we added an ethylene scavenger, PAs, a PA inhibitor, and compounds that affect ethylene biosynthesis and activity to the regeneration medium. Regeneration frequencies increased in response to treatment with ethylene inhibitors (aminoethoxyvinylglycine and silver thiosulfate) and an ethylene scavenger (mercury perchlorate). Treatment with the ethylene precursor 1-aminocyclopropane-1-carboxylic acid reduced the regeneration frequency, increased callus formation, and increased ethylene levels; similar results were obtained in response to treatment with the PA inhibitor methylglyoxal-bis(guanylhydrazone). By contrast, treatment with PAs (particularly spermidine and spermine) decreased ethylene levels, increased the regeneration frequency, and increased shoot bud formation. These results suggest a coordinated regulation of ethylene and polyamines because the suppression of ethylene levels using ethylene inhibitors, polyamines, or mercury perchlorate increased the in vitro regeneration frequency and morphogenic responses of Capsicum annuum L.

Microgravity effects on different stages of higher plant life cycle and completion of the seed‐to‐seed cycle

Human inhabitation of Space requires the efficient realisation of crop cultivation in bioregenerative life-support systems (BLSS). It is well known that plants can grow under Space conditions; however, perturbations of many biological phenomena have been highlighted due to the effect of altered gravity and its possible interactions with other factors. The mechanisms priming plant responses to Space factors, as well as the consequences of such alterations on crop productivity, have not been completely elucidated. These perturbations can occur at different stages of plant life and are potentially responsible for failure of the completion of the seed-to-seed cycle. After brief consideration of the main constraints found in the most recent experiments aiming to produce seeds in Space, we focus on two developmental phases in which the plant life cycle can be interrupted more easily than in others also on Earth. The first regards seedling development and establishment; we discuss reasons for slow development at the seedling stage that often occurs under microgravity conditions and can reduce successful establishment. The second stage comprises gametogenesis and pollination; we focus on male gamete formation, also identifying potential constraints to subsequent fertilisation. We finally highlight how similar alterations at cytological level can not only be common to different processes occurring at different life stages, but can be primed by different stress factors; such alterations can be interpreted within the model of 'stress-induced morphogenic response' (SIMR). We conclude by suggesting that a systematic analysis of all growth and reproductive phases during the plant life cycle is needed to optimise resource use in plant-based BLSS.

An improved in vitro encapsulation protocol, biochemical analysis and genetic integrity using DNA based molecular markers in regenerated plants of Withania somnifera L

Non-embrogenic, synthetic seeds were produced by encapsulating nodal segments (containing axillary buds) of Withania somnifera L. in calcium alginate hydrogel containing Murashige and Skoog (MS) medium. A 3% sodium-alginate with 100mM calcium cloride found to be the optimum concentration for the production of uniform synthetic seeds. Effect of different treatments (M1–M5), i.e. MS medium containing different concentrations of cytokinins (0.5, 1.0, 2.5, 5.0 & 10.0μM) along with optimal level of auxins NAA (0.5μM) on in vitro morphogenic response of synthetic seeds was evaluated. The maximum frequency (86.2%) of conversion of encapsulated beads into plantlets was achieved on MS (M3) medium containing 6-benzyladenine, BA (2.5μM) and α-naphthalene acetic acid, NAA (0.5μM) after 4 weeks of culture. Rooting in plantlets was achieved on 1/2MS+NAA (0.5μM). Plantlets obtained from stored synthetic seeds were hardened, acclimatized and established in field, where they grew well without any detectable malformation. Significant enhancement in the pigment contents (chlorophyll, carotenoids and net photosynthetic rates) with an increase in acclimatization days may be attributed to chlorophyll biosynthesis. Activities of antioxidant enzymes i.e. superoxide dismutase, catalase and peroxidase activity) were significantly increased which suggests their preventive role in membrane oxidation and damage to biological molecules. Also, an enhanced level of lipid peroxidation, as indicated by MDA content, a sensitive diagnostic index of oxidative injury clearly indicating its positive determining role in combating oxidative stress during acclimatization of plantlets. The generated RAPD and ISSR profiles from regenerated plantlets with mother plant were monomorphic which confirms the genetic stability among the clones. This synthetic seed technology could possibly paves the way for the conservation, short-term storage, germplasm exchange with potential storability and limited quarantine restrictions.

L-methionine influences in vitro root regeneration, total chlorophyll, total carbohydrate and endogenous proline content in the sweet cherry rootstock MxM 14 (Prunus avium × Prunus mahaleb)

In the present study, the effects of indole-3-butyric acid (IBA) separately and simultaneously with l-methionine on the morphogenic and biochemical responses in the cherry rootstock MxM 14 (Prunus avium×Prunus mahaleb), from shoot tip explants, were investigated. Best rooting results were achieved with 1mg/L IBA plus 0.5mg/L l-methionine. The incorporation of the amino acid in the Murashige and Skoog (MS) medium, greatly enhanced the promotive effect of the auxin on rooting, exhibiting a synergistic action. The supplementetion of explants with l-methionine did not influence their shoot length, for each IBA treatment. On the other hand, l-methionine when combined with the highest IBA concentration (2mg/L) had an inhibitory effect on shoot fresh and dry weight. l-methionine significantly affected chlorophyll, carbohydrate and proline concentration. The maximum leaf chlorophyll concentration was recorded with 2mg/L IBA plus 1mg/L l-methionine. IBA alone or combined with l-methionine doubled the leaf carbohydrate concentration. Moreover, proline concentration in leaves was significantly increased with low (0.5mg/L) l-methionine, irrespective of IBA concentration. In roots, the combined effect of 0.5mg/L IBA with 0.5mg/L l-methionine and the 1mg/L IBA plus 1mg/L l-methionine treatment led to elevated levels of endogenous proline and total carbohydrates, respectively, in comparison to the control. Taking into account the different parameters examined, chlorophyll and carbohydrates appeared as the most accurate biochemical markers of the rooting process in the MxM 14 rootstock.

In Vitro Propagation and Synthetic Seeds Production: An Efficient Methods for Stevia rebaudiana Bertoni

Germplasm can be effectively stored in the form of synthetic seeds. Shoot tips obtained from in vitro shoot cultures of Stevia rebaudiana Bertoni were encapsulated in 4 % calcium alginate. The present work discussed the role of components of culture medium on morphogenic response of Stevia rebaudiana Bertoni encapsulated buds to various MS strengths and sugar alcohol (Mannitol or sorbitol) in different concentrations for long term storage. Germination ability of the synthetic seeds was investigated. Shoots were regenerated from nodal explants of Stevia through axillary shoot proliferation. The induction of multiple shoots from nodal segments was the highest in MS medium supplemented with 1.0 mg l−1 BA. Maximum shoot formation was obtained with fructose at 20 and 40 g l−1 fructose, while with fructose at 20 g l−1 gave the highest leaves number/explants). The longest shoot length obtained with sucrose and fructose than other sugar. Different media type (NN and WPM) were suitable for best shoot number of Stevia, leaf number and shoot length than other media. Growth of shoot was increased and observed in capsules stored for 5 weeks on MS than other MS strengths. The growth of capsules dependent on mineral concentration and storage time. The most suitable conversions of capsules was using 0.05 M mannitol after 6 weeks from storage of synthetic seeds of Stevia. For rooting, when Stevia shoots cultured on MS medium supplemented with IAA, at 0.2 mg l−1 resulted in the maximum number of roots/explant while, IBA at 1.0 and 2.0 mg l−1 resulted in longest root/plant and gave the same length of root.

LEAFY COTYLEDON2 (LEC2) promotes embryogenic induction in somatic tissues of Arabidopsis, via YUCCA-mediated auxin biosynthesis

The LEAFY COTYLEDON2 (LEC2) transcription factor with a plant-specific B3 domain plays a central role in zygotic and somatic embryogenesis (SE). LEC2 overexpression induced in planta leads to spontaneous somatic embryo formation, but impairs the embryogenic response of explants cultured in vitro under auxin treatment. The auxin-related functions of LEC2 appear during SE induction, and the aim of the present study was to gain further insights into this phenomenon. To this end, the effect of LEC2 overexpression on the morphogenic responses of Arabidopsis explants cultured in vitro under different auxin treatments was evaluated. The expression profiles of the auxin biosynthesis genes were analysed in embryogenic cultures with respect to LEC2 activity. The results showed that LEC2 overexpression severely modifies the requirement of cultured explants for an exogenous auxin concentration at a level that is effective in SE induction and suggested an increase in the auxin content in 35S::LEC2-GR transgenic explants. The assumption of an LEC2 promoted increase in endogenous auxin in cultured explants was further supported by the expression profiling of the genes involved in auxin biosynthesis. The analysis indicated that YUCCAs and TAA1, working in the IPA-YUC auxin biosynthesis pathway, are associated with SE induction, and that the expression of three YUCCA genes (YUC1, YUC4 and YUC10) is associated with LEC2 activity. The results also suggest that the IAOx-mediated auxin biosynthesis pathway involving ATR1/MYB34 and CYP79B2 does not seem to be involved in SE induction. We conclude that de novo auxin production via the tryptophan-dependent IPA-YUC auxin biosynthesis pathway is implicated in SE induction, and that LEC2 plays a key role in this mechanism.Electronic supplementary materialThe online version of this article (doi:10.1007/s00425-013-1892-2) contains supplementary material, which is available to authorized users.

Effects of exogenous l-arginine on in vitro rooting, chlorophyll, carbohydrate, and proline concentrations in the sweet cherry rootstock M × M 14 (Prunus avium L. × Prunus mahaleb L.)

The effects of indole-3-butyric acid (IBA) alone and in combination with l-arginine on the morphogenic and biochemical responses in shoot tip explants of the cherry rootstock M × M 14 (Prunus avium × Prunus mahaleb) were examined. The maximum root number per rooted explant (16), root fresh (FW) and dry (DW) weights, as well as the rooting percentage (100 %) were recorded when 2 mg l−1 IBA (alone) were applied. Including the lowest IBA concentration (0.5 mg l−1) with the lowest and highest l-arginine concentrations (0.5 and 2 mg l−1, respectively) resulted in the greatest root length. The maximum leaf chlorophyll concentration and shoot length of the initial explant were recorded when 0.5 mg l−1 IBA plus 2 mg l−1l-arginine were applied. In addition, l-arginine in combination with IBA (1 and 2 mg l−1) was found to suppress shoot FW and DW. On the other hand, l-arginine enhanced the promoting effect of IBA on both root length and leaf chlorophyll concentration. The carbohydrate and proline concentrations in leaves were not significantly altered with the application of IBA alone or in combination with l-arginine. On the other hand, the carbohydrate and proline concentrations in roots were decreased with the application of 1 and 2 mg l−1 IBA with l-arginine, resulting in the suppression of the promoting effects of IBA. It is clear from the findings that l-arginine has a direct effect on the in vitro rooting of M × M 14 explants, is involved in the function of the photosythetic apparatus, influences leaf chlorophyll content, participates in carbohydrate biosynthesis and metabolism, and is involved in proline accumulation both in leaves and roots.

Correlation between hormonal homeostasis and morphogenic responses in Arabidopsis thaliana seedlings growing in a Cd/Cu/Zn multi‐pollution context

To date, almost no information is available in roots and shoots of the model plant Arabidopsis thaliana about the hierarchic relationship between metal accumulation, phytohormone levels, and glutathione/phytochelatin content, and how this relation affects root development. For this purpose, specific concentrations of cadmium, copper and zinc, alone or in triple combination, were supplied for 12 days to in vitro growing seedlings. The accumulation of these metals was measured in roots and shoots, and a significant competition in metal uptake was observed. Microscopic analyses revealed that root morphology was affected by metal exposure, and that the levels of trans-zeatin riboside, dihydrozeatin riboside, indole-3-acetic acid and the auxin/cytokinin ratio varied accordingly. By contrast, under metal treatments, minor modifications in gibberellic acid and abscisic acid levels occurred. Real-time polymerase chain reaction analysis of some genes involved in auxin and cytokinin synthesis (e.g. AtNIT in roots and AtIPT in shoots) showed on average a metal up-regulated transcription. The production of thiol-peptides was induced by all the metals, alone or in combination, and the expression of the genes involved in thiol-peptide synthesis (AtGSH1, AtGSH2, AtPCS1 and AtPCS2) was not stimulated by the metals, suggesting a full post-transcriptional control. Results show that the Cd/Cu/Zn-induced changes in root morphology are caused by a hormonal unbalance, mainly governed by the auxin/cytokinin ratio.

The mitochondrial glutathione peroxidase GPX3 is essential for H2O2 homeostasis and root and shoot development in rice

Glutathione (GSH) peroxidases (GPXs: EC 1.11.1.9 and EC1.11.1.12) are non-heme thiol peroxidases that catalyze the reduction of H2O2 or organic hydroperoxides to water, and they have been identified in almost all kingdoms of life. The rice glutathione peroxidase (OsGPX) gene family is comprised of 5 members spread throughout a range of sub cellular compartments. The OsGPX gene family is induced in response to exogenous H2O2 and cold stress. In contrast, they are down regulated in response to drought and UV-B light treatments. Transgenic rice plants have been generated that lack mitochondrial OsGPX3. These GPX3s plants showed shorter roots and shoots compared to non-transformed (NT) plants, and higher amounts of H2O2 mitochondrial release were observed in the roots of these plants cultivated under normal conditions. This accumulation of H2O2 is positively associated with shorter root length in GPX3s plants compared to NT ones. Moreover, GPX3 promoter analysis indicated that it is mainly expressed in root tissue. These results suggest that silencing the mitochondrial OsGPX3 gene impairs normal plant development and leads to a stress-induced morphogenic response via H2O2 accumulation.

Production, storability and morphogenic response of alginate encapsulated axillary meristems and genetic fidelity evaluation of in vitro regenerated Ceropegia bulbosa: A pharmaceutically important threatened plant species

In vitro multiple shoot production was achieved in Ceropegia bulbosa var. bulbosa. Nodal segments obtained from mature plant were used as an explant and their morphogenetic potential was tested on, Murashige and Skoog's medium containing cytokinins alone or in combination with auxins. Application of 8.88μM, 6-benzyladenine was most effective for shoot regeneration and resulted in an average of 9.7 shoots/explant. Nodal segments obtained from in vitro proliferated shoots of C. bulbosa, were encapsulated in calcium alginate beads for large-scale clonal propagation, short-term conservation, germplasm exchange and distribution. Maximum percent response for conversion of encapsulated nodal segments into plantlets was 100%. Encapsulated nodal segments could be stored at low temperature (4°C) up to 60 days with a survival frequency of 50.7%. Morphologically identical plants generated in vitro were genetically identical when analyzed using random decamer primers (RAPD).

Synthetic Seed Technology for Encapsulation and Regrowth of in vitro Derived Stevia rebaudiana Bert. nodal Segments

Synthetic seed technology is an alternative to traditional micropropagation for production and delivery of cloned plantlets. Regrowth of nodal segments of Stevia rebaudiana Bert. after encapsulation in calcium alginate matrix was evaluated. These nodal segments were derived from in vitro cultured shoots. Encapsulation was performed with 5% sodium alginate and 50 mM CaCl2. Morphogenic response and percentage of conversion into plantlets of encapsulated buds on various planting media was tested. Encapsulated explants exhibited the best regrowth and conversion frequency on Murashige and Skoog medium supplemented with TDZ (0.5µM) under in vitro conditions with 85% survival rate. Plantlets regenerated from the encapsulated explants were hardened off and successfully transferred to soil. Acknowledgements: The work was supported by United States Department of Agriculture Agricultural Research Service Specific Cooperative Agreement No. 58 – 6408 – 02 – 1-612.