Diversity and potential application of endophytic bacteria in ginger.

Here, 248 endophytic bacterial strains were isolated to assess the distribution and population diversity of endophytic bacteria in ginger plants. A total of 10.4 x 10(4) to 20.2 x 10(4) CFU/g fresh weight endophytic bacteria of different growth stages were isolated. Maximum bacterium numbers were obtained at the seedling stage. A total of 107 functional strains were screened, including 17 antibacterial strains and 90 indole acetic acid-producing strains. Based on 16S rDNA sequence restriction fragment length polymorphism and 16S rDNA sequences, these 107 strains were mapped and grouped into 16 genera. Bacillus and Pseudomonas were the dominant genera; however, the bacteria belonged to a tremendous range of genera, with the highest species richness being observed at the seedling stage. Sixteen strains exhibited antimicrobial activity against Pythium myriotylum Drechsler, while 7 strains exhibited antimicrobial activity against Phyllosticta zingiberi Hori. Bacillus was the dominant antibacterial strain. Pseudomonas fluorescens, B. megaterium, and Enterobacter ludwigii produced remarkably high levels of IAA. Only a few endophytic bacterial strains were inhibited in fresh ginger juice. Most of these strains were present during seedling stage, including Roseateles depolymerans, Chryseobacterium taiwanense, E. ludwigii, Agrobacterium larrymoorei, P. fluorescens, and Bacillus amyloliquefaciens. This study indicates that the community of endophytic bacteria in ginger changes with the synthesis of antibacterial substances.

Cytological and embryological studies on apospory in Bothriochloa ischaemum L.

Cytological and embryological studies on apomictic species Bothriochloa ischaemum L. were carried out. Our studies revealed that the chromosome number of its root apical cells was 40, indicating that it was a tetraploid cytotype. During the stage of microsporogenesis, meiosis seemed irregular, as the pairing chromosome number of microspore mother cell was more than 20. It was often found that some chromosomes did not assemble in the equatorial plane or moved to the two poles of the cell, a few laggards were seen. Multiporate pollens (22.3%) were often observed. The studies showed that a high frequency (87.8%) of 1-3 or more aposporous embryo sacs developed in one ovule of the species. The mature aposporous sac was usually characterized by an egg cell and one polar nucleus. The egg cell could develop spontaneously into a large proembryo (100-200 microm) mass prior to anthesis. When several aposporous sacs occurred in the same ovule, usually 2 aposporous sacs were involved in pseudogamy and developed into separate endosperm masses in the same ovary. In the low frequency of mature seed, 13.5% twin-embryo seedlings could be obtained after mature seeds germinated.

Ultrafast and large third-order nonlinear optical properties of CdS nanocrystals in polymeric film.

We report the ultrafast and large third-order nonlinear optical properties of CdS nanocrystals (NCs) embedded in a polymeric film. The CdS NCs of 2 nm radius are synthesized by an ion-exchange method and highly concentrated in the two layers near the surfaces of the polymeric film. The two-photon absorption coefficient and the optical Kerr coefficient are measured with laser pulses of 250 fs duration at 800 nm wavelength. The one-photon and two-photon figures of merit are determined to be 3.1 and 1.3, respectively, at irradiance of 2 GW/cm(2). The observed nonlinearities have a recovery time of approximately 1 ps. The two-photon-generated free carrier effects have also been observed and discussed. These results demonstrate that CdS NCs embedded in polymeric film are a promising candidate for optical switching applications.

Preparation of uniform titanium dioxide (TiO2) polystyrene-based composite particles using the glass membrane emulsification process with a subsequent suspension polymerization.

Uniform titanium dioxide (TiO(2))-polystyrene-based composite particles were prepared using the glass membrane emulsification process followed by a subsequent suspension polymerization. The oil phase, consisting of anatase TiO(2) fine powder, monomers, methyl laurate as the hydrophobic additive, Disperbyk-180 and the poly(styrene-co-2-ethyl hexylacrylate) were emulsified through the membrane pores into the aqueous phase containing stabilizers to form a (solid-in-oil)-in-water (S/O/W) emulsion of monomer droplets. The suspension polymerization was carried out at 343 K for 24 h under a nitrogen atmosphere. An SPG membrane with a pore size of 5.25 microm was employed and 20-25 microm TiO(2)-polystyrene based composite particles were obtained depending on the composition of polymerizing oil phase. The effects of the co-monomer, 2-ethylhexyl acrylate and the cross-linking agent, divinyl benzene on the dispersion stability of TiO(2) in the oil phase, the surface feature of the particle and the encapsulation loading were investigated in this study. The membrane emulsification process was capable of preparing the composite particles with approximately 5 wt% of TiO(2) encapsulated, which accounts for with at least 85 wt% of TiO(2) in the oil phase.

Performance of titanium dioxide microcapsules as a photo-oxidation catalyst for decolourization of methylene blue.

The performance of anatase type titanium dioxide (TiO(2)) encapsulated with styrenic copolymers via the solvent evaporation or suspension polymerization process was investigated as a photocatalyst for the decolourization of methylene blue (MB, 3,4-bis(dimethylamino)-phenothiazine-5-thionium chloride) in the aqueous phase. The TiO(2) microcapsules, loaded with 4-8 wt% TiO(2), were dispersed in a MB aqueous solution containing an adequate amount of hydroperoxide as an oxygen source, and the mixture was exposed to 365 nm UV light. The decolourization of MB proceeded according to the first order of the MB concentration. The apparent rate constant, defined based on the unit weight of loaded TiO(2), depended on the initial concentration of MB. The capsule walls, composed of cross-linked and/or uncross-linked poly(styrene-co-2-ethylhexyl acrylate), favoured the adsorption of MB on the capsule surface and promoted the decomposition. The observations of the cross-section of microcapsules by transmission electron microscopy (TEM) showed similar morphology of microcapsules regardless of preparation method; a thin layer of hydrophilic TiO(2) particles being localized near the particle surface. The sustainability of the microcapsules was also proved by doing experiments successively for 7 days. Smaller size microcapsules were favoured for the decomposition of MB, and the rate constant increased with the surface area of microcapsules in unit volume of the reaction mixture. Although large size microcapsules with uncross-linked polymer wall had a disadvantage of breaking under high shear agitation, those prepared with cross-linked polymer wall by suspension polymerization still remained effective after the sustainability test for 7 days.

Uniform titanium dioxide (TiO(2)) microcapsules prepared by glass membrane emulsification with subsequent solvent evaporation.

Anatase-type titanium dioxide (TiO(2)) was encapsulated using an Shirasu porous glass (SPG) membrane emulsification technique and followed by solvent evaporation. The oil phase, consisting of fine#10; powder of anatase TiO(2), Disperbyk-180, the hydrophobic oil phase additive, and polymer wall solution, was pushed through the membrane pores into the aqueous phase of poly(vinyl alcohol) and sodium dodecyl sulfate to form the solid-in-oil-in water, (S/O)/W, emulsion droplets. Three types of styrene-based copolymer poly(styrene-co-acrylic acid) (PS-AA), poly(styrene-co-2-ethyl hexyl acrylate) (PS-2EHA) and poly(styrene-co-dimethyl aminoethylmethacrylate) (PS-DMAEMA) were used as an encapsulating shell. Uniform droplets were successfully obtained by modifying the oil phase using methyl laurate or hexadecanol as the oil phase additive, together with carefully monitoring the emulsification flow rate during the emulsification. The (S/O)/W emulsion was gently stirred in a sealed reactor, and evacuation of solvent started under moderate heating with increasing a vacuum intensity. Those uniform-sized TiO(2) microcapsules revealed fine porous morphologies on their surfaces as a result of a mild phase separation induced from the addition of the oil phase additive. The encapsulation efficiency was influenced by the stability of TiO(2) in the oil phase, the polymer wall employed, and the operational control of the glass membrane emulsification process. The membrane emulsification process could prepare the TiO(2) microcapsules with about approximately 6-8.5 wt% of encapsulation loadings. PS-AA and PS-2EHA copolymers provided better encapsulation efficiency compared to PS-DMAEMA. SPG membranes with 1.42, 2.8, 5.25, 7.0, or 9.5 microm were employed and 2-20 microm microcapsules were subsequently obtained.

Magnetite (Fe3O4) microcapsules prepared using a glass membrane and solvent removal.

Fine magnetite powders dispersed in polymer solution were encapsulated from an oil-in-water emulsion prepared by an emulsification process employing a porous glass membrane and subsequent evaporation of the solvent. Styrene-based copolymers were dissolved in a magnetic fluid, and then continuously pushed through the pores of glass membrane into the aqueous phase, which had dissolved polyvinyl alcohol (PVA) and sodium dodecyl sulphate (SDS) as a mixed stabilizer. P(styrene-co-acrylic acid) (PS-AA), P(stryrene-co-butyl acrylate) (PS-BA) and styrene-butadiene rubber (SBR) were dissolved in the specially ordered magnetite fluid (25 wt% magnetite dispersed in toluene) separately or as a mixture, and uniform droplets suspending the magnetic particles were obtained. After the evaporation of toluene, PS-AA capsules retained a spherical shape and uniformity, whereas PS-AA/PS-BA capsules revealed a creased surface and broader size distribution. The microcapsules entrapped 30-40 wt% of magnetite, and the encapsulation yield of magnetite was 20-40%. Glass membranes with 9.5, 5.25 and 1.42 microm pore size were employed and 5-40 microm microcapsules were obtained depending on the pore size. When magnetite suspended in chloroform was used, magnetite capsules with broader size distributions were obtained because of the sticking of the droplets to the membrane wall. The advantage of the membrane emulsification which provides uniform sized droplets was lost.

Selective G-protein regulation of neuronal calcium channels.

We examined the properties and regulation of Ca channels resulting from the expression of human alpha1B and alpha1E subunits stably expressed in KEK293 cells. The ancillary subunits beta1B and alpha2/delta were also stably expressed in these cell lines. Ca currents in alpha1B-expressing cells had the properties of N-type currents. Ca currents in cells expressing alpha1E exhibited a novel profile that was similar to the properties of the "R type" Ca current. Introduction of GTP-gamma-S into alpha1B cells greatly enhanced the extent of prepulse facilitation of the Ca current, whereas it had only a very small effect in alpha1E-expressing cells. Activation of somatostatin receptors endogenous to HEK293 cells or kappa opioid receptors, expressed in the cells after transfection, inhibited Ca currents in alpha1B-expressing cells. This inhibition was blocked by pertussis toxin and was partially relieved by a depolarizing prepulse. In contrast, no inhibitory effects were noted in cells expressing alpha1E channels under the same circumstances. HEK293 cells normally contained G-proteins from all of the four major families. Inhibition of Ca currents by kappa agonists in alpha1B-expressing cells was enhanced slightly by the cotransfection of several G-protein alpha subunits. kappa agonists, however, had no effect in alpha1E-containing cells, even after overexpression of different G-protein alpha-subunits. In summary, these results demonstrate that there is a large difference in the susceptibility of alpha1B- and alpha1E-based Ca channels to regulation by G-proteins. This is so despite the fact that the two types of Ca channels show substantial similarities in their primary sequences.

Functional expression of an epitope-tagged G protein-coupled K+ channel (GIRK1).

An epitope-tagged form of an inwardly rectifying and G protein-coupled K+ channel (GIRK1-cp) was expressed at high levels in transfected mammalian cells. Immunoblot analysis of transfected human embryonic kidney cells (HEK293) and mouse insulinoma cells (beta TC3) revealed several GIRK1-cp polypeptides, including the major 59-kDa band, corresponding to the predicted mass of the GIRK1 polypeptide plus the epitope tag. Immunohistochemical staining using two anti-tag antibodies showed abundant immunoreactive material, which was predominantly concentrated in the perinuclear area in both transfected cell types. While functional GIRK1-cp message was present in poly(A)+ RNA prepared from HEK293 cells expressing GIRK1-cp protein, appropriate K+ currents could not be detected. In contrast, whole cell recordings made directly from transfected beta TC3 cells expressing GIRK1-cp revealed inwardly rectifying, pertussis toxin-sensitive currents activated by norepinephrine and galanin. Single channel recordings in excised patches of beta TC3 cells expressing GIRK1-cp showed rectifying K+ currents when activated by 50 microM guanosine 5'-O-(thiotriphosphate), with a slope conductance of 39.1 +/- 1.0 picosiemens. This is the first report of stable heterologous expression of a functional G protein-coupled K+ channel in mammalian cells. The activity of an epitope-tagged channel in insulinoma cells demonstrates the utility of this system for further biochemical and biophysical analyses of G protein-K+ channel interactions.

kappa-Opioid receptor activates an inwardly rectifying K+ channel by a G protein-linked mechanism: coexpression in Xenopus oocytes.

cRNAs encoding the kappa-opioid receptor and an inwardly rectifying, G protein-coupled, K+ channel were coinjected into Xenopus oocytes. The effects of kappa-opioid receptor agonists and antagonists on the membrane currents in these oocytes were studied using the two-electrode voltage-clamp technique. The kappa-opioid receptor agonists U69593 and dynorphin A induced a concentration-dependent inward current (EC50 of approximately 0.3 microM and approximately 30 nM, respectively) after coinjection of both cRNAs, whereas the mu-opioid receptor agonist [D-Ala2,N-MePhe4,Gly5-ol]enkephalin (10 microM) and the delta-opioid receptor agonist [D-Pen2,5]enkephalin (1 microM) had no effect. The agonist-induced inward current was reversible upon washing out of the agonists and was inhibited in the presence of the K+ channel blocker Ba2+ (0.1 mM). The specific kappa-opioid receptor antagonist norbinaltorphimine (0.1 microM) and the nonspecific opioid receptor antagonist naloxone (1 microM) abolished the agonist-induced currents. Furthermore, the agonist-induced currents exhibited rapid desensitization in the continuous presence of the agonists or after repeated application. Preincubation of the coinjected oocytes with pertussis toxin (400 ng/ml for 3 days of 1.5 microgram/ml for 24 hr) abolished most of the agonist-induced activation of the inwardly rectifying K+ current. We therefore conclude that specific stimulation of the kappa-opioid receptor can activate the inwardly rectifying K+ channel through a pertussis toxin-sensitive G protein.

[Study on biosynthesis of indigo involving transferring naphthalene plasmid DNA from Pseudomonas to E. coli].

Indigo is one of the brilliant blue dyes, which was used to be extracted from plants, but now synthesized chemically. Indigo is also produced by bacteria. In recent years, Ensley, B. D. (1983) and Mermod, N. (1986) reported a pathway producing indigo of bacteria. We are currently studying indigo formation by bacteria. Using Pseudomonas sp. S13 harboring naphthalene degradation plasmid as donor and E. coli as recipient, conjugates and transformants with the plasmid were obtained. The conjugates and transformants were not only able to grow in medium with naphthalene as the sole carbon and energy source, but also to synthesize indigo in Lennox medium. The conjugates and transformants were grown in Lennox medium at 30 degrees C for 48 hr then resulted in synthesis of indigo. The production of indigo is increased in the presence of tryptophan or indole. Indigo formation was enhanced if the bacteria was grown in a medium supplemented with either 0.1% of naphthalene or 1% of salicylic acid. The present work opens a field for the synthesis of dyes by microbes and it might shed light on the potential use in controlling environmental pollution by engineering bacteria.