High Agar Concentration Research Articles

Direct shoot regeneration from cotyledonary node of Uraria picta (Jacq.) Desv. ex DC., an important plant of dashmula drugs, and assessment of genetic fidelity, metabolic profiling, and anti-diabetic activity

Uraria picta root is used in the polyherbal product ‘Dashmula’. Its exploitation for formulation preparation has depleted its availability, leading to medicine adulteration. Direct shoot regeneration from the cotyledonary node holds promise as a source of raw material. This study aimed to develop a regeneration protocol for U. picta and validate its genetic and metabolic fidelity. The seeds of U. picta showed low germination rates, prolonged dormancy, and poor viability. Root exploitation in the wild poses a threat to its availability in nature. Seedling’s derived cotyledonary nodes cultured on B5 medium supplemented with BAP (0.5–3 mg L−1), Kinetin (0.5–3 mg L−1), Thidiazuron (0.01–1 mg L−1), and meta-Topolin (0.1–4 mg L1). To address hyperhydricity in regenerated shoots, cotyledonary nodes were cultured on high-agar concentration media. Microshoots were exposed to IBA solution (50–800 mg L−1) pulse treatment for rooting. Tissue-cultured plants genetic fidelity was assessed using ISSR and SCoT markers, while metabolic fidelity was studied with HRMS. The chlorophyll content, antioxidant, and antidiabetic activity of micropropagated plants were evaluated. The highest shoot regeneration frequency, with a maximum of 6.57±0.278 shoots per explant, was achieved using 2 mg L−1meta-Topolin. The shoots were elongated, had expanded leaves, and were hyperhydrated. BAP (2 mg L−1) induced a maximum of 9.83±0.333 shoot buds per explant. BAP caused explant browning, profuse callus formation, dwarfing, and hyperhydric shoots. Hyperhydricity was alleviated with a higher agar concentration (1 %). IBA (400 mg L−1) induced a maximum of 2.18±0.090 roots per shoot and a root length of 9.23±0.033 cm. Tissue-cultured and mother plants exhibited clonal fidelity, similar metabolite and chlorophyll content, strong antioxidant activity, and equal efficacy for inhibiting α-amylase and α-glucosidase. This method can propagate elite clones of U. picta and offer its improvement via genetic transformation.

Gliding motility of a uranium-tolerant Bacteroidetes bacterium Chryseobacterium sp. strain PMSZPI: insights into the architecture of spreading colonies.

Uranium-tolerant soil bacterium Chryseobacterium sp. strain PMSZPI moved over solid agar surfaces by gliding motility thereby forming spreading colonies which is a hallmark of members of Bacteroidetes phylum. PMSZPI genome harboured orthologs of all the gld and spr genes considered as core bacteroidetes gliding motility genes of which gldK, gldL, gldM and gldN were co-transcribed. Here, we present the intriguing interplay between gliding motility and cellular organization in PMSZPI spreading colonies. While nutrient deficiency enhanced colony spreading, high agar concentrations and presence of motility inhibitor like 5-hydroxyindole reduced the spreading. A detailed in situ structural analysis of spreading colonies revealed closely packed cells forming multiple layers at centre of colony while the edges showed clusters of cells periodically arranged in hexagonal lattices interconnected with each other. The cell migration within colony was visualized as branched structures wherein the cells were buried within extracellular matrix. PMSZPI colonies exhibited strong iridescence possibly as a result of periodicity within the cell population achieved through gliding motility. Presence of uranium reduced motility and iridescence and induced biofilm formation. The coordinated study of gliding motility and iridescence apparently influenced by uranium provides unique insights into the lifestyle of PMSZPI residing in uranium enriched environment.

Optimizing Transformation Frequency of Cryptococcus neoformans and Cryptococcus gattii Using Agrobacterium tumefaciens.

The transformation of Cryptococcus spp. by Agrobacterium tumefaciens has proven to be a useful genetic tool. A number of factors affect transformation frequency. These factors include acetosyringone concentration, bacterial cell to yeast cell ratio, cell wall damage, and agar concentration. Agar concentration was found to have a significant effect on the transformant number as transformants increased with agar concentration across all four serotypes. When infection time points were tested, higher agar concentrations were found to result in an earlier transfer of the Ti-plasmid to the yeast cell, with the earliest transformant appearing two h after A. tumefaciens contact with yeast cells. These results demonstrate that A. tumefaciens transformation efficiency can be affected by a variety of factors and continued investigation of these factors can lead to improvements in specific A. tumefaciens/fungus transformation systems.

Effects of culture medium composition and PEG on hyperhydricity in Dendrobium officinale

Hyperhydricity is a physiological disorder during plant tissue culture that seriously affects regeneration and micropropagation. In this study, Dendrobium officinale plantlets were cultured on solid Murashige and Skoog (MS) medium supplemented with plant growth regulators and various concentrations of sucrose, agar, and polyethylene glycol (PEG)-6000 to explore the effect of osmotic stress on hyperhydricity. The results show that low concentrations of sucrose or agar, as well as PEG-6000 at various concentrations, significantly increase the hyperhydric rate of D. officinale, whereas high concentrations of sucrose or agar did not. Furthermore, high concentrations of PEG-6000 significantly increase total water content, free-water content, relative electrical conductivity, and peroxidase (POD) activity of D. officinale plantlets, whereas they significantly decrease bound-water content, proline content, soluble protein content, soluble sugar content, and superoxide dismutase (SOD) activity. These results indicate that PEG-6000 disrupts the antioxidant system and water metabolism in D. officinale plantlets, as well as increases cell membrane permeability, which might be the key factors for the occurrence of hyperhydricity in this species.

Establishment and initial development in vitro of Yellow Ipe

Micropropagation protocol success clearly depends on the establishment and initial development in vitro . Following steps can only be performed after obtaining of aseptic cultures endowed with good plant vigor. This study aimed to evaluate different culture medium and agar concentrations in the establishment and early development in vitro yellow ipe. For both experiments the source of explants were shoot apical segments, obtained in vitro seed germination. For the first experiment was used a ½ WPM (diluted to 50% normal salt concentration) and different concentrations of agarose (4, 6, 8 and 10 g L -1 ). For the experiment of culture nutrient medium treatments were: control (agar and water); MS medium; ½ and ¼ MS; WPM; ½ and ¼ WPM. After 30 days of culture were performed evaluations. For the agar concentration experiment evaluated the percentage of contamination, establishment, in vitro root formation and callus, number of roots and leaves. For the experiment of culture medium evaluated the percentage of establishment and in vitro formation of roots, number of roots and leaves. With increasing concentrations of agar added to the culture medium, it was found that there is a significant increase in the formation and in the number of roots per plant yellow ipe. The WPM and the diluted ½ and ¼ provided greater root formation and with better training, and more training leaves. We conclude that higher agar concentration, the lower the callus formation and the WPM and their dilutions are effective in establishing and initial in vitro development of yellow ipe.

Sucrose release from agar gels and sensory perceived sweetness

Sweetness resulting from sucrose released from agar gels of varying concentrations of agar (0.5, 1.0, and 1.5% w/w) and sucrose (10%–50% w/w) was evaluated. The intensity of sweetness as perceived by subjects was investigated during consumption of the gels. A time–intensity method was applied: the maximum intensity and area under the time–intensity curve were higher in gels with lower concentrations of agar as expected from previous studies. The intensity of sweetness as estimated by the area under the time–intensity curve increased with sucrose concentration, but the maximum intensity tended to reach a plateau above a certain sucrose concentration. The manifestation of the maximum sweetness was delayed with increasing concentrations of sucrose and agar. The total duration of the perception of sweetness was longer in gels with higher sucrose concentrations but did not depend on the concentration of agar in the range examined. Instrumental characterization of these gels was also conducted. Young's modulus, fracture stress, fracture strain, and energy obtained from a conventional uniaxial compression test of sample gels increased with sucrose concentration and the modulus, stress, and energy were higher in gels with higher agar concentration. Although mechanical characteristics such as Young's modulus, fracture stress, strain, and energy, increased with increasing sucrose concentration, the sweetness intensity tended to a plateau; therefore, physiological and psychological factors experienced during oral processing should be taken into account to understand the perceived sweetness intensity.

Induction and development of anther-derived gametic embryos in cucumber (Cucumis sativus L.) by optimizing the macronutrient and agar concentrations in culture medium

Anther culture response of a cucumber cultivar (Beta Alpha) and three Iranian landraces of cucumber (Basmenj, Isfahani, and Korki) on MS culture media with an altered macronutrient strength (half, full, 1.5-fold, and double-strength) or various concentrations of agar (0, 3.5, 7, and 14 g L-1) was evaluated in two separate experiments. The effect of different concentrations of agar on the development of callus-derived embryos was also assessed. The androgenic potentials of various cucumbers were significantly (P < 0.01) influenced by the different macronutrient strengths and agar concentrations, although anther culture responses could vary according to the cucumber type. A doubled strength of macronutrients in culture medium resulted in the highest percentage of callogenesis (100%) in Isfahani cucumber, while use of full strength of macronutrients produced the highest percentage of embryogenic calli (83.33%) in the mentioned landrace compared to other tested forms. The maximum number of gametic embryos (1.26 and 1.23 embryos/anther) differentiated from Isfahani anther cultures on full- and half-strength MS medium, respectively. In the second experiment, the highest percentages of callogenesis were obtained in MS culture media solidified with 14 g L-1 and 7 g L-1 agar for Basmenj, with 3.5 g L-1 agar or liquid medium for Isfahani, with 7 g L-1 agar for Beta Alpha, and with 14 g L-1 agar for Korki. Culture media solidified with 7 g L-1 agar produced the highest percentage of embryogenic calli compared to other treatments. Anther culture of Korki cucumber on culture medium with 14 g L-1 agar produced the maximum number of embryos per anther compared to other cucumbers. Improved production of mature somatic embryos (torpedo and cotyledonary embryos) was achieved with higher agar concentrations (7 and 14 g L-1) for the cucumber cultivar and all landraces.

Improved in vitro propagation of Rose cultivars against varying concentrations of carbon source, agar and plant growth regulators

The present study was conducted to improve upon the micropropagation protocol of Rose cultivars by modification of the MS medium through variations in sucrose and agar concentrations thereby enhancing the shoot multiplication and rooting efficiency of the nodal explants. High sucrose concentrations and low agar concentrations favoured shoot mutiplication during the in vitro stages. Enhanced multiplcation and growth was observed on sub culturing the mother explants with regenerated shoots on fresh MS medium containing sucrose (3.5%, w/v) and agar (0.6%, w/v), supplemented with lower concentrations cytokinin combination of BAP and KN (2.5 mgl-1+1.5 mgl-1) respectively. Half strength MS medium containing sucrose (2.0%, w/v) and agar (0.3%, w/v) with NAA and BAP (2.0 mgl-1+0.5 mgl-1) in combination was most effective for rooting.

A Novel Cellular Autoaggregative Developmentally CRP Regulated Behaviour Generates Massively Chondrule-Like Formations over Surface of Old &amp;lt;i&amp;gt;Escherichia coli&amp;lt;/i&amp;gt; K-12 Macrocolony Biofilms

How Escherichia coli bacteria develop a particular colonial, 3-D biofilm morphological pattern is still a poorly understood process. Recently, we reported a new E. coli K-12 morphotype exhibited by old macrocolonies described as volcano-like. The formative developmental process of this morphotype has been presented as a suitable experimental model for the study of 3D patterning in macrocolony biofilms. Here, we report the optical microscopy observations and genetic analysis that have unveiled the existence of a novel autoaggregative behaviour which generates massive lumpiness over the surface of the volcano-like macrocolonies. These lumpy formations are generated by the autoaggregation and strong interaction of tightly packed bacterial cells in structures with a chondrule-like appearance which give the colony’s surface its characteristic microscopic lumpy phenotype. Furthermore, they exhibit different levels of maturation from the edge to the center of the colony. Hence, its generation appears to follow a spatiotemporal program of development during the macrocolony’s morphogenesis. Interestingly, the agar’s hardness influences the morphology exhibited by these formations, with high agar concentration (1.5%, 15 g/L) suppressing its development. This new auto-aggregative E. coli’s behaviour does not require the activity of the biofilm master regulator CsgD, the adhesiveness of flagella, pili type 1, adhesin Ag43, β-1,6-N-acetyl-D-glucosamine polymer-PGA, cellulose or colanic acid, but it is under glucose repression and the control of cAMP receptor protein (CRP). The possible physiological role of these chondrule-like formations in the adaptability of the colony to different stressful environmental conditions is discussed.

The MARVEL domain protein Nce102 regulates actin organization and invasive growth of Candida albicans.

ABSTRACTInvasive growth of the fungal pathogen Candida albicans into tissues promotes disseminated infections in humans. The plasma membrane is essential for pathogenesis because this important barrier mediates morphogenesis and invasive growth, as well as secretion of virulence factors, cell wall synthesis, nutrient import, and other processes. Previous studies showed that the Sur7 tetraspan protein that localizes to MCC (membrane compartment occupied by Can1)/eisosome subdomains of the plasma membrane regulates a broad range of key functions, including cell wall synthesis, morphogenesis, and resistance to copper. Therefore, a distinct tetraspan protein found in MCC/eisosomes, Nce102, was investigated. Nce102 belongs to the MARVEL domain protein family, which is implicated in regulating membrane structure and function. Deletion of NCE102 did not cause the broad defects seen in sur7Δ cells. Instead, the nce102Δ mutant displayed a unique phenotype in that it was defective in forming hyphae and invading low concentrations of agar but could invade well in higher agar concentrations. This phenotype was likely due to a defect in actin organization that was observed by phalloidin staining. In support of this, the invasive growth defect of a bni1Δ mutant that mislocalizes actin due to lack of the Bni1 formin was also reversed at high agar concentrations. This suggests that a denser matrix provides a signal that compensates for the actin defects. The nce102Δ mutant displayed decreased virulence and formed abnormal hyphae in mice. These studies identify novel ways that Nce102 and the physical environment surrounding C. albicans regulate morphogenesis and pathogenesis.

Evaluation of Agar-Bound Microparticulate Diet as Alternative Food in Abalone Hatchery: Effects of Agar Concentrations and Feeding Frequencies

The performance of an agar-bound microparticulate diet (A-MPD) was evaluated on feeding postlarval abalone Haliotis asinina, focusing on the effects of agar concentrations and feeding frequencies. Larval abalone, obtained from the Southeast Asian Fisheries Development Center, Aquaculture Department hatchery, were reared in 60-L flow-through tanks with UV-filtered seawater. They were fed 1,200 mg A-MPD bound with either 5.0 mg/mL agar solution, 7.5 mg/mL agar solution, 10.0 mg/mL agar solution, and 12.5 mg/mL agar solution, or a natural diet consisting of diatoms at different feeding frequencies (daily, every other day, or every 2 d) starting at day 5. A 5 × 3 factorial experiment in a completely randomized design tested the effects of various treatments on postlarval settlement and survival after days 15 and 90. Scheffé's postcomparison test determined differences among treatments means. Postlarval settlement and survival were not significantly different in diets bound with higher agar concentrations and tested in 3 feeding frequencies. At lower levels of agar incorporation in diets, however, settlement and survival counts became significantly higher on daily feeding. Postlarval settlement and survival were significantly highest with abalone fed a diet bound with 7.5 mg/mL agar solution on a daily feeding frequency. Average percent weight loss in the feed was higher with lower levels of agar incorporation. Average particle size of both A-MPD and diatoms was 4–5 µm. Crude protein content of A-MPD was 42.7%; that of diatoms was 14.9%. A-MPD may be used as alternative food in abalone hatcheries with the incorporation of 7.5 mg/mL agar solution fed daily to abalone.

Diffusion of Methylene Blue in Phantoms of Agar Using a Photoacoustic Technique

In this work, the kinetics of diffusion of methylene blue in agar aqueous solution is studied using a photoacoustic technique. Two agar phantoms solutions in water with a relation of mass/volume of 0.01% and 0.05% were analyzed. The study was performed using a modified Rosencwaig photoacoustic cell that is enclosed by transparent windows, on both sides. The sample is deposited directly on top of the upper window. A red light beam, at a fixed modulation frequency, is sent through the lower window illuminating the sample and inducing the photoacoustic effect inside the closed chamber of the cell. At the beginning of the experiment, a droplet of 100μL of agar solution is deposited; afterwards, the signal stabilizes, and 10μL of methylene blue aqueous solution (0.0125 g · mL−1) is added to the surface of the agar. During the first seconds of the experiment, the photoacoustic signal amplitude increases followed by a gradual and long decay. Results for modulation frequencies in the range from 10Hz to 80Hz for both agar concentrations are presented. A simple theoretical approach is presented to analyze the experimental data. It is demonstrated that the kinetics of the process can be parameterized as a function of the changes of an effective optical absorption coefficient. From these results, the characteristic time, in which the dye diffusion process stabilizes, is obtained. It is found that this time is larger for samples with a higher agar concentration. These differences provide important results for biomedical sciences in which agar gels are used as phantoms resembling some of the properties of living organs and tissues.

Investigations into Protein Crystallization in the Presence of a Strong Magnetic Field

A new strategy is proposed for batch crystallization of proteins in solution-growth or gel-growth by using the batch method inside capillary tubes applying magnetic fields. Four proteins with differing proportions of α-helices and β-sheets and crystallized in five different crystallographic space groups are studied, allowing an analysis of the anisotropy of the diamagnetic susceptibility of the peptide bond as well as the polarity of the space groups in the presence of a strong magnetic field of 11.75 T. The crystal quality is shown to be improved by using a strong magnetic field to orient protein molecules, and gel-growth (high concentrations of agar) to control the transport phenomena as well as crystal growth. Some advantages to increase the crystal quality for crystals from marginal conditions for X-ray diffraction, and disadvantages of the use of solution- and gel-growth (low concentration of agar) in magnetic fields, and their plausible applications to high resolution X-ray crystallography are discussed.

Effect of agar concentration on the matric potential of glycerol agar media and the germination and growth of xerophilic and non-xerophilic fungi

The concept of water activity ( a w) does not differentiate between water status resulting from the interaction of water with solutes, and that from interaction of water with matrices, which is termed matric potential ( Ψ m). This study reports the effect of agar concentration (1.5, 3.0, 4.5, 6.0, and 7.5%, w/w) on matric potential of glycerol agar media (GA) and the germination and growth of xerophilic fungi ( Eurotium herbariorum and E. rubrum) and non-xerophilic fungi ( Aspergillus niger, A. flavus, and Penicillium roqueforti) on GA with or without sorbic acid (0.1–0.4%, w/w) at 0.90 a w and 0.95 a w. The matric potential of GA decreased when the agar concentration increased from 1.5 to 7.5%. When the agar concentration increased at each a w, the radial growth rate of the xerophilic fungi generally increased but the biomass density (biomass per unit area) decreased, whereas the radial growth rate of the non-xerophilic fungi generally decreased but the biomass density was unchanged. In the absence of sorbic acid, the time to germination of each species was similar for all agar concentrations. In the presence of sorbic acid, the time to germination of some species was significantly longer at higher agar concentrations (4.5%–7.5%) than 1.5% agar. This study demonstrated the inhibition effect on germination and growth of non-xerophilic fungi and xerophilic fungi by decreased matric potential resulting from increased agar concentrations, and the different responses of non-xerophilic fungi and xerophilic fungi to water stress from solutes and matrices. The concept of matric potential may be useful in food microbiology to provide a better understanding of fungal growth in complex food matrices.

Swarming and complex pattern formation in Paenibacillus vortex studied by imaging and tracking cells

BackgroundSwarming motility allows microorganisms to move rapidly over surfaces. The Gram-positive bacterium Paenibacillus vortex exhibits advanced cooperative motility on agar plates resulting in intricate colonial patterns with geometries that are highly sensitive to the environment. The cellular mechanisms that underpin the complex multicellular organization of such a simple organism are not well understood.ResultsSwarming by P. vortex was studied by real-time light microscopy, by in situ scanning electron microscopy and by tracking the spread of antibiotic-resistant cells within antibiotic-sensitive colonies. When swarming, P. vortex was found to be peritrichously flagellated. Swarming by the curved cells of P. vortex occurred on an extremely wide range of media and agar concentrations (0.3 to 2.2% w/v). At high agar concentrations (> 1% w/v) rotating colonies formed that could be detached from the main mass of cells by withdrawal of cells into the latter. On lower percentage agars, cells moved in an extended network composed of interconnected "snakes" with short-term collision avoidance and sensitivity to extracts from swarming cells. P. vortex formed single Petri dish-wide "supercolonies" with a colony-wide exchange of motile cells. Swarming cells were coupled by rapidly forming, reversible and non-rigid connections to form a loose raft, apparently connected via flagella. Inhibitors of swarming (p-Nitrophenylglycerol and Congo Red) were identified. Mitomycin C was used to trigger filamentation without inhibiting growth or swarming; this facilitated dissection of the detail of swarming. Mitomycin C treatment resulted in malcoordinated swarming and abortive side branch formation and a strong tendency by a subpopulation of the cells to form minimal rotating aggregates of only a few cells.ConclusionP. vortex creates complex macroscopic colonies within which there is considerable reflux and movement and interaction of cells. Cell shape, flagellation, the aversion of cell masses to fuse and temporary connections between proximate cells to form rafts were all features of the swarming and rotation of cell aggregates. Vigorous vortex formation was social, i.e. required > 1 cell. This is the first detailed examination of the swarming behaviour of this bacterium at the cellular level.

The RssAB Two-Component Signal Transduction System in Serratia marcescens Regulates Swarming Motility and Cell Envelope Architecture in Response to Exogenous Saturated Fatty Acids

Serratia marcescens swarms at 30 degrees C but not at 37 degrees C on a nutrient-rich (LB) agar surface. Mini-Tn5 mutagenesis of S. marcescens CH-1 yielded a mutant (WC100) that swarms not only vigorously at 37 degrees C but also earlier and faster than the parent strain swarms at 30 degrees C. Analysis of this mutant revealed that the transposon was inserted into a gene (rssA) predicted to encode a bacterial two-component signal transduction sensor kinase, upstream of which a potential response regulator gene (rssB) was located. rssA and rssB insertion-deletion mutants were constructed through homologous recombination, and the two mutants exhibited similar swarming phenotypes on LB swarming agar, in which swarming not only occurred at 37 degrees C but also initiated at a lower cell density, on a surface with a higher agar concentration, and more rapidly than the swarming of the parent strain at 30 degrees C. Both mutants also exhibited increased hemolysin activity and altered cell surface topologies compared with the parent CH-1 strain. Temperature and certain saturated fatty acids (SFAs) were found to negatively regulate S. marcescens swarming via the action of RssA-RssB. Analysis of the fatty acid profiles of the parent and the rssA and rssB mutants grown at 30 degrees C or 37 degrees C and under different nutrition conditions revealed a relationship between cellular fatty acid composition and swarming phenotypes. The cellular fatty acid profile was also observed to be affected by RssA and RssB. SFA-dependent inhibition of swarming was also observed in Proteus mirabilis, suggesting that either SFAs per se or the modulation of cellular fatty acid composition and hence homeostasis of membrane fluidity may be a conserved mechanism for regulating swarming motility in gram-negative bacteria.

Tissue culture and regeneration of thallus from callus of Gelidiella acerosa (Gelidiaies, Rhodophyta)

The tissue culture of an economically important red alga Gelidiella acerosa (Gelidiales, Rhodophyta) included preparation of axenic material, culture of explants, subculture of excised callus and regeneration of de novo plants from callus in the laboratory. Sequential treatment of explants with sterile seawater consisting of 0.1% liquid detergent for 10 min, 2% betadine (with 0.5% w/v available iodine) for 5 min and 3.5% broad-spectrum antibiotic mixture with nystatin for 2 days enabled yields as high as 90% of viable and axenic explants. A prolific and rapid growth of filamentous callus on explants was observed on cut surfaces during the first month of culture. The highest level of callus induction occurred in Provasoli enriched seawater (PES) medium solidified with 1.5% agar incubated at 20–22°C and a photon flux density of 5 μmol photons m−2 s−1 with a 12: 12 light-dark photoperiod. Up to 90% of the explants cultured at 5 μmol photons m−2 s−1 produced callus, whereas at 30 and 70 μmol photons m−2 s−1, 70% and 9% produced callus, respectively. The explant culture medium with 0.5% agar content stimulated bud development in all explants, whereas higher agar concentrations (0.8%, 1.0%, 1.5%, 2.0% and 3.0%) resulted in a filamentous type of callus growth. Addition of the plant growth regulators naphthalene acetic acid and indole 3-acetic acid (auxins), and benzyl amino purine and kinetin (cytokinins) and different organic carbon supplements (glycerol, sucrose, sorbitol and mannitol) to the culture medium had no effect on callus growth or induction rate. All carbon supplies at 0.5 and 1.0 M concentration showed an inhibitory effect and most of the explants perished gradually after 2 weeks in culture. The callus mass with bud or shoot developments continued to grow when transferred to semisolid PES medium (0.2% agar w/v) on a rotary shaker. In 4 months, these shoots gave rise to 2–3 cm long plantlets of G. acerosa. The tissue-cultured Gelidiella germlings were successfully grown into full plants in the field on coral stones in 6 months.

Effect of Gelling Agent on Growth and Development of Ceratozamia hildae Somatic Embryos

I. Abstract Individual somatic proembryos of Ceratozamia hildae were exposed to media that differed only in gelling agent utilized. Five different gelling agents were compared in the first experiment: Bacto agar, Agargel, Gel-Gro, Phytagar, and TC agar. In addition, the effect of agar was examined at two levels. Growth, proliferation, and development were assessed. The lower level of agar did not support good somatic proembryo growth. Agargel and the high agar concentration produced cultures with good proliferation. Proembryos exposed to Phytagar, Gel-Gro, and TC agar had the highest proliferation rates. Overall, Gel-Gro was considered the best gelling agent tested. The three concentrations of Gel-Gro used in the second experiment were 2, 4, and 6 g·l−1, with the lowest concentration representing the control, the recommended concentration. As gelling agent concentration increased, so did mortality; however, the highest Gel-Gro concentration also produced the highest numbers of good-quality, mature somatic...

培地中の糖および寒天濃度，光強度がサツマイモ培養小植物体の成長および炭素吸収に及ぼす影響

The effects of sucrose and agar concentration in the media, and light intensity on growth and carbon absorption were observed using the plantlets of sweet potato (Ipomoea batatas (L.) Lam.), The stem cuttings with one leaf dissected from in vitro cultured plantlets were cultured under 12 degrees of combinations of three sugar levels (5, 20, and 45 gl-1) and two agar levels (5 and 10 gl-1) in medium, and two levels of PPFD (50 and 100 μmol m-2 s-1) maintained at an air temperature of 25±1°C and a carbon dioxide concentration of 1, 000 μmol mol-1. The dry weight of the plantlets increased as the sugar concentration and/or light intensity increased. Under high agar concentration, the shoot growth declined, but the root growth accelerated. Absorbed molar quantity of carbon dioxide was more in high concentration of agar or high light intensity. The plantlets which was inhibited the sugar absorption by the high agar concentration seemed to depend more on CO2 as a carbon source. It may be suitable that the media contain high sugar and agar concentration in early stage for heterotrophic or mixotrophic growth of the plantlets, and then CO2 concentration and light intensity are gradually raised for their mixotrophic or autotrophic growth according to the status of the plantlet.

Anatomical analysis of Saccharomyces cerevisiae stalk-like structures reveals spatial organization and cell specialization.

Recently we reported an unusual multicellular organization in yeast that we termed stalk-like structures. These structures are tall (0.5 to 3 cm long) and narrow (1 to 3 mm in diameter). They are formed in response to UV radiation of cultures spread on high agar concentrations. Here we present an anatomical analysis of the stalks. Microscopic inspection of cross sections taken from stalks revealed that stalks are composed of an inner core in which cells are dense and vital and a layer of cells (four to six rows) that surrounds the core. This outer layer is physically separated from the core and contains many dead cells. The outer layer may form a protective shell for the core cells. Through electron microscopy analysis we observed three types of cells within the stalk population: (i) cells containing many unusual vesicles, which might be undergoing some kind of cell death; (ii) cells containing spores (usually one or two spores only); and (iii) familiar rounded cells. We suggest that stalk cells are not only spatially organized but may undergo processes that induce a certain degree of cell specialization. We also show that high agar concentration alone, although not sufficient to induce stalk formation, induces dramatic changes in a colony's morphology. Most striking among the agar effects is the induction of growth into the agar, forming peg-like structures. Colonies grown on 4% agar or higher are reminiscent of stalks in some aspects. The agar concentration effects are mediated in part by the Ras pathway and are related to the invasive-growth phenomenon.