黑蒴定芽诱发增殖的初步研究

黑蒴(Melasma arvense)为云南省民族民间珍稀药材,现已成为云南天然药物开发的热点药材种类.但目前在云南已濒临灭绝,急需对其进行资源保护和人工繁育.该研究在筛选黑蒴不同器官培养的基础上,以茎枝为外植体,依次考察了不同基本培养基、不同种类细胞分裂素和质量浓度、不同天然有机添加物对定芽(顶芽和侧芽)萌发增殖及生长的影响.结果表明:以黑蒴带节茎枝为外植体,将其剪成2~3 cm带节嫩茎枝,经严格的外植物体消毒程序处理后,接种到1/2MS+6-BA 0.5 mg·L-1+Pt 50 g·L-1+Bn 80 g·L-1+25 g·L-1蔗糖+7 g·L-1琼脂,pH值5.8的培养基上,接种后放置在光照强度1500~2000 lx,光照10 h·d-1,温度22℃下培养,接种后8~10 d开始萌发,40 d单茎平均增殖数达8.1,主茎40 d生长长度74.2 mm,显示出极好的诱发增殖生长效果.该研究结果初步建立了黑蒴嫰茎枝诱发定芽及增殖培养体系,为云南珍稀民族药黑蒴的资源保护、人工栽培和可持续利用提供了科学依据.     

Multiplication of juvenile black wattle by microcuttings

The influence of mineral formulation, growth regulators and activated charcoal on micropropagation of juvenile black wattle (Acacia mearnsii) was studied. Nodal segments of one-month-old seedlings were used as starting material. After 30 to 45 days, they developed into shoots, which were divided into microcuttings and transferred to fresh media. The addition of 2 g l⁻¹ of activated charcoal to basal medium (3/4 strength MS salts and MS vitamins) improved the multiplication phase, reducing leaf chlorosis and raising the percentage of elongated shoots. The multiplication rate varied between 1.7 and 3 every month. Rooting percentage too was higher in the presence of charcoal, even when auxin was not added to the culture medium. The addition of 8.87 μM of benzyladenine and/or 1.44 or 2.88 μM of gibberellic acid did not influence significantly the multiplication, nor modifications in the FeSO₄, MnSO₄, CaCl₂ content of the medium. The system described allows the multiplication, elongation and rooting of black wattle in one step. This revised version was published online in June 2006 with corrections to the Cover Date.     

Diseased leaf lyophilization: a method for long-term prevention of loss of virulence in phytopathogenic bacteria

A technique for long-term preservation of phytopathogenic bacteria is described. It is based on selective multiplication of the bacterial pathogen in host tissue, disinfection of leaf surfaces to reduce contamination, lyophilization of leaves, and storage under dry conditions at - 80°C. With this technique, the pathogenicity of large numbers of the pathogens Pseudomonas syringae pv. tomato and Xanthomonas campestris pv. vesicatoria was maintained for 4 years.     

Establishment and multiplication of red clover plants by in vitro shoot tip culture

A procedure is described for the routine establishment and multiplication of red clover, Trifolium pratense L., shoot tips which should be applicable to a wide genotypic background. The addition of CO₂ to the culture vial or use of polypropylene closures enhanced shoot multiplication at high levels of benzyladenine (BA). Horizontal orientation of crown buds resulted in more efficient multiplication. Culture of nodes from flowering stems was unsuccessful. The cytokinin BA was most effective for shoot multiplication at 2.0 mg/l with maximum shoot production by four weeks. A comparison of several genotypic sources revealed a 10-fold range in response to the multiplication medium with no differences observed among agronomic type or ploidy level. An additional study revealed that multiplication ability of a genotype can be determined after the second subculture since multiplication ability does not change during repeated subculture.     

Changes with background in the linear model of the transient visual system

There is evidence that the transient channel of temporal human vision behaves as a linear filter for small excursions around a steady background level. The linear filter characteristics depend on the background level. From experimentally obtained impulse responses of the transient channel the linear filter can be modelled and parametrized. This has been done for two different background levels. The two sets of estimated parameters at these two levels show a shift in the parameters which can be described by a single multiplication factor. This result was extrapolated to arbitrary background levels by postulating that each change in background level can be described by a multiplication factor. This leads to an assumption on the variation of the parameters of the linear filter of the transient channel with changes in the background level. This assumption is tested by simulating the system for different parameter sets of the linear filter. The simulations give a good agreement with experimental data on threshold-versus-duration curves and de Lange curves. The (minor) quantitative differences in simulations and experimental data can be explained.     

A new monoclonal antibody enzyme-linked immunosorbent assay to measure in vitro multiplication of the microsporidium Encephalitozoon intestinalis

Microsporidia are obligate intracellular eukaryotic parasites that infect a wide range of hosts, including invertebrates and vertebrates. Microsporidia have emerged as important opportunistic pathogens of humans with the onset of the AIDS pandemic. The potential impact of these infections in human pathology has required the development of antiparasitic strategies, based on the search for molecules having an effect on the development and/or the multiplication of microsporidia. This creates a demand for a simple and reliable in vitro technique for measuring the multiplication of microsporidia. We developed a new monoclonal antibody (MAb) enzyme-linked immunosorbent assay (ELISA) technique and measured the growth of Encephalitozoon intestinalis in an in vitro culturing system using this method. The monoclonal antibody is specific for a coat protein of E. intestinalis sporogonic stages produced in parasitophorous vacuole. An anti-mouse antibody labeled with peroxidase was used as conjugate. This ELISA is a suitable, specific and semiquantitative technique for measuring the spread of E. intestinalis. It is easy to perform and required 5 h from start to end. A good correlation was observed when the ELISA data were compared with the manual microscopic counts of parasitophorous vacuoles obtained after immunofluorescent assay (IFA). Moreover, the ELISA method proved more accurate than the immunofluorescent assay. In summary, the ELISA system described in this study provides a simple reliable assay for measuring the spread of microsporidia in vitro and may prove valuable for the screening of putative interesting antimicrosporidial compounds.     

SERUM FRACTIONATION AND THE EFFECTS OF BOVINE SERUM FRACTIONS ON HUMAN CELLS GROWN IN A CHEMICALLY DEFINED MEDIUM

Serum has been fractionated by curtain electrophoresis using carboxymethyl cellulose dissolved in sodium bicarbonate electrolyte. Various fractions were produced from bovine serum and added to replicate cultures of Chang's endoepithelial cells and HeLa cells grown in a chemically defined medium. The effects of each of the various fractions on the appearance of the cultures and on cell multiplication were studied. Three different fractions were obtained and two were subjected to further purification. One fraction associated with albumin promoted survival, attachment, and flattening as well as cell multiplication. A second fraction associated with the alpha globulins promoted survival and multiplication of some cells. A third fraction caused cells to aggregate and form free floating clumps. An adequate chemically defined medium for continuous growth of human cells was used throughout the study. The response of cells to alterations in their environment which simulated some of the effects produced by serum fractions is described.     

Establishment and multiplication of red clover plants by in vitro shoot tip culture

A procedure is described for the routine establishment and multiplication of red clover, Trifolium pratense L., shoot tips which should be applicable to a wide genotypic background. The addition of CO₂ to the culture vial or use of polypropylene closures enhanced shoot multiplication at high levels of benzyladenine (BA). Horizontal orientation of crown buds resulted in more efficient multiplication. Culture of nodes from flowering stems was unsuccessful. The cytokinin BA was most effective for shoot multiplication at 2.0 mg/l with maximum shoot production by four weeks. A comparison of several genotypic sources revealed a 10-fold range in response to the multiplication medium with no differences observed among agronomic type or ploidy level. An additional study revealed that multiplication ability of a genotype can be determined after the second subculture since multiplication ability does not change during repeated subculture.Contribution from the Plant Cell Culture Centre, University of Guelph, Department of Crop Science, Guelph, Ontario, Canada N1G 2W1     

Continuousin vitro multiplication of shoot buds of Norway spruce (Picea abies L.) by intermittent application of growth regulators

A culture system was developed which allows continuous production of adventitious buds. Segments of seedlings germinated on media supplemented with different growth regulator combinations were used as explants. Cultivation was carried out in two phases, which alternate permanently: (I) without and (II) with growth regulators. Thus it was possible to establish meristematic tissue cultures from 5–10% of the seedlings tested. They have produced buds now for over 3 years with multiplication rates of about 1.5 within 5–6 weeks.     

Multiplication of human diploid fibroblasts in a synthetic medium supplemented with EGF, insulin, and dexamethasone

Substantial multiplication of human diploid fibroblasts (HDF) has been obtained in medium MCDB 108 supplemented with epidermal growth factor (EGF), insulin, and dexamethasone (DEX). Growth rate is somewhat slower than in serum-supplemented medium. However, large wellformed colonies can be obtained in 14 days, and sequential monolayer subculture is possible up to a total of about ten population doublings. A basal medium that has been optimized specifically for HDF is essential for such multiplication. In addition, polylysine-coated culture surfaces, low temperature trypsinization, and careful removal or neutralization of residual trypsin are also needed. The culture system contains no deliberately-added undefined components, and is chemically defined except for possible roles of contaminants in the materials that are used for its preparation.     

Compartmentalization of self-reproducing machineries: Multiplication of microsystems with self-instructing polymerization of amino acids

A theoretical model is presented for the self-instructing polymerization of free amino acids which proceeds inside microsystems which are phase-separated from the solution of thermal polyamino acids. It is shown theoretically that a compartmentalized microsystem fixes inside itself only the process with a faster macromolecular multiplication as time passes, even if the catalytic polymerization alone could spontaneously decrease the corresponding reaction rate. The compartmentalized machinery of macromolecular multiplication cannot reach its stationary state. The machinery is inevitably multiplied and alternates with those with either faster rates of macromolecular multiplication or slower rates of macromolecular degradation during their time development. These results are based upon the dynamic process that any material system acts by itself so as to remove any flow disequilibrium, that is, to maintain the continuity of material flow.     

Molecular characterization of the genetic integrity of wheat (Triticum aestivum L.) germplasm after long-term maintenance

The genetic identity of eight wheat (Triticum aestivum L.) accessions maintained in the Gatersleben genebank and regenerated up to 24 times was studied by using wheat microsatellite markers (WMS). It was demonstrated that WMS can be used to analyze bulks of seeds stored more than 50 years in a seed reference collection at room temperature. No contamination due to foreign pollen or incorrect handling during the multiplication cycles was discovered. For one accession (TRI 4599) genetic drift was observed, whereas for TRI 249 a heterogenous situation for two markers was maintained over the years. We were able to show that microsatellites can be used as a simple and reliable marker system for the verification of the integrity and genetic stability of genebank accessions. Received: 29 March 1999 / Accepted: 22 June 1999     

Synthesis of Some Purine Carbocyclic Isosteres of 2′,3′-Dideoxy-3′-C-Hydroxymethyl Nucleosides as Potential Inhibitors of HIV

Abstract

The synthesis of some enantiomerically pure carbocyclic 2′,3′-dideoxy-3′-C-hydroxymethyl derivatives of adenine, inosine and guanine is described. The Mitsunobu reaction was used in the coupling procedure giving exclusively N⁹-coupling. The nucleosides were tested for inhibition of HIV multiplication in vitro and were found to be inactive in the assay.     

The effect of feed salinity on the biofouling dynamics of seawater desalination

A persistent cell labeling dye and a novel microbial counting method were used to explore the effects of salinity on a microbial population in a reverse osmosis (RO) desalination system, and these clearly distinguished microbial cell multiplication from cell adherence. The results indicated that microbial multiplication is more active at the front of a seawater RO pressure vessel, while adhesion dominates the back of the vessel. A severe reduction in RO permeate flux and total dissolved solid (TDS) rejection were detected at low salinity, attributed to marked cell multiplication and release of extracellular polymeric substances, whilst a relatively stable flux was observed at medium and high salinity. The results from PCR-DGGE revealed the variation in microbial species distribution on the membrane with salinity. The results imply the critical role of membrane modification in biofouling mitigation in the desalination process.     

Control of potato cyst nematode (Globodera pallida) by host plant resistance and nematicide

Seven trials conducted over four years on sites naturally infested with the white potato cyst nematode established that potato clones bred for resistance to Globodera pallida allowed significantly less nematode multiplication than conventional cultivars under field conditions. Nematode multiplication was inversely related to initial infestation level. The nematicide, aldicarb, significantly reduced nematode multiplication. However, nematode multiplication on nematicide treated susceptible cultivars was greater than on untreated partially resistant clones, indicating that resistance may offer more effective control of G. pallida than chemical treatment. Integration of host plant resistance and nematicide treatment is discussed.     

Landscape structure and locust swarming: a satellite's eye view

Desert locust Schistocerca gregaria outbreaks consistently start in the same places, suggesting that certain landscapes are particularly favourable for outbreaking. Outbreaks are generated by multiplication, concentration and gregarisation of locust populations. Previous research has shown how small-scale vegetation patterns in desert ecosystems influence locust gregarisation; the present study examines the effects of large-scale landscape structure on locust multiplication and concentration. NOAA/AVHRR satellite imagery was used to relate abundance and spatial distribution of resources at the landscape scale to the historical record of locust outbreaks. Threshold NDVI values were investigated to define what constitutes 'resource' for locusts. The first part of the study showed that abundance and spatial distribution of resource were not sufficient to distinguish between outbreak and non-outbreak areas in the western part of the locust distribution area. Thus, outbreak danger zones cannot be identified by landscape structure at this spatial resolution. The second analysis investigated spatio-temporal patterns of vegetation growth in two locust breeding areas with very different landscape structure; in both cases, the patterns differed significantly between outbreaking and non-outbreaking years. In Mauritania, a flat homogeneous desert landscape, both resource abundance and fragmentation were higher in outbreaking years. On the Red Sea coast, a fragmented landscape, resource spatial distribution was consistent between years, and abundance alone was a significant predictor of outbreaking. High resource abundance promotes locust multiplication, and contraction of resource into small patches increases locust concentration; these two mechanisms explain how landscape structure influences locust outbreaking.     

Efficient procedure for grapevine embryogenic suspension establishment and plant regeneration: role of conditioned medium for cell proliferation

An efficient system for the establishment and multiplication of highly prolific embryogenic cell cultures of grapevine (Vitis sp.) was developed. Using anther-derived pro-embryogenic masses as starting material, cell suspensions of different grapevine cultivars (Tempranillo, Cabernet-Sauvignon) and rootstocks (Kober 125 AA, Kober 5 BB, 110 Richter) were initiated in liquid medium containing NOA (1.0 mg l(-1)) and BAP (0.25 mg l(-1)) as growth regulators. Conditioned medium was recovered and utilised for establishing new, highly totipotent cell cultures. The suspensions obtained, showed embryogenic competence resulting in somatic embryo induction and subsequent plant regeneration. In this study, a simplified establishment procedure for grapevine embryogenic cell suspension allowing the fast multiplication of embryogenic material is described. Evidence for the promoting effect of the protein fraction derived from conditioned medium, on cell proliferation was found. In bioassays, addition of ss-D: -GlcY affect cell proliferation suggesting that arabinogalactan proteins are required for growth processes in grapevine cell cultures.     

From growth arrest to growth suppression.

Since the introduction of the cell cycle concept two approaches to study growth regulation of cells have been proposed. One claims that cells are naturally quiescent, requiring a stimulatory encouter with growth factors for induction of cell division. The other considers cellular multiplication as the natural steady-state; cessation of multiplication is thus a restriction imposed on the system. In the latter case emphasis is mainly on the signals involved in arrest of multiplication. This Prospect focuses on specific events occurring in mammalian cells at growth arrest, senescence, and terminal differentiation, specifically emphasizing the growth inhibitory factors, tumor suppressor genes, and other signals for growth suppression.     

Density-Dependent Nematode Seasonal Multiplication Rates and Overwinter Survivorship: A Critical Point Model

Nematode multiplication rates Pf/Pi and overwinter survivorship (Pi2/Pfl) for Meloidogyne incognita were both adequately described by negative exponential models, indicating density dependence in each case. Density dependence of the multiplication rates is mediated by resource limitation and host damage; in survivorship rates it may be mediated by limitation of stored reserves or prevalence of antagonists. Parameters of multiplication rate models were crop specific and varied with host status and environmental suitability. Maximum multiplication rates (a) of nearly 1,000 were measured for tomatoes. Equilibrium densities were sensitive to tolerance of the nematode by the crop. Overwinter survival rates varied among locations where cultural practices and length of infestation time differed.     

Naringenin is a novel inhibitor of Dictyostelium cell proliferation and cell migration

Naringenin is a flavanone compound that alters critical cellular processes such as cell multiplication, glucose uptake, and mitochondrial activity. In this study, we used the social amoeba, Dictyostelium discoideum, as a model system for examining the cellular processes and signaling pathways affected by naringenin. We found that naringenin inhibited Dictyostelium cell division in a dose-dependent manner (IC(50) approximately 20 microM). Assays of Dictyostelium chemotaxis and multicellular development revealed that naringenin possesses a previously unrecognized ability to suppress amoeboid cell motility. We also found that naringenin, which is known to inhibit phosphatidylinositol 3-kinase activity, had no apparent effect on phosphatidylinositol 3,4,5-trisphosphate synthesis in live Dictyostelium cells; suggesting that this compound suppresses cell growth and migration via alternative signaling pathways. In another context, the discoveries described here highlight the value of using the Dictyostelium model system for identifying and characterizing the mechanisms by which naringenin, and related compounds, exert their effects on eukaryotic cells.