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  • Immunocytochemistry  (43)
  • Saccharomyces cerevisiae  (43)
  • rice  (37)
  • maize  (36)
  • Springer  (158)
  • Nature Publishing Group
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  • 2010-2014
  • 1990-1994  (158)
  • 1970-1974
  • 1990  (158)
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  • 2010-2014
  • 1990-1994  (158)
  • 1970-1974
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  • 1
    Electronic Resource
    Electronic Resource
    Springer
    Entomologia experimentalis et applicata 55 (1990), S. 285-294 
    ISSN: 1570-7458
    Keywords: Graminella nigrifrons ; maize ; oats ; johnsongrass ; development ; fecundity ; host suitability
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Description / Table of Contents: Résumé La dynamique des populations (durée de développement de l'œuf à l'adulte, poids et taille des adultes, fécondité) de G. nigrifrons Forbes (Homop. Cicadellidae) a été étudiée au laboratoire à 5 températures sur plantules de maïs (Zea mays L.), avoine (Avena sativa L.) et sorgho vivace (Shorgum halepense (L.) Pers.). Sur les 3 plantes, les mâles se développent en moyenne 1,2 j plus vite que les femelles. Les relations entre vitesse de développement et température ont été déterminées en utilisant à la fois un modèle linéaire et le modèle biophysique à 2 paramètres de Sharpe & DeMichele (1977). Les températures plus basses donnent des adultes des 2 sexes plus gros et plus lourds. Moins de G. nigrifrons se sont développés sur la graminée vivace que sur les 2 graminées annuelles à la température la plus élevée (30°C), tandis qu'à la température la plus basse (18°C) moins de cicadelles se sont développées sur les graminées annuelles. La température semble jouer un rôle significatif en déterminant l'adéquation des plantes comme hôtes convenant au développement de G. nigrifrons. Le potentiel de ponte de cette cicadelle avait été sous-estimé par les étudies précédentes.
    Notes: Abstract Population dynamics of the blackfaced leafhopper, Graminella nigrifrons (Forbes) (Homoptera: Cicadellidae), was studied at five temperatures (18, 21, 24, 27, & 30°C) in the laboratory on seedling maize (Zea mays L.), oats (Avena sativa L.), and the perennial johnsongrass (Sorghum halepense (L.) Pers.). Effects of temperature and host plant on egg to adult mean development time, adult size and weight, and fecundity were determined. Leafhoppers on all three hosts developed fastest at the highest temperature tested (21.3 days), and slowest at the lowest temperature tested (73.2 days). The duration from first to last adult eclosion was shortest at 30°C, (11.5 days) and longest at 18°C (43 days). The sex ratio of males to females did not differ from 1:1, but males developed an average of 1.2 days faster than females on all three hosts. Mean percent development/day ranged from 1.4% at 18°C to 4.7% at 30°C. The relationship of this development rate and temperature was determined using both a linear model and a variable parameter biophysical model. Based on these models, the developmental threshold is estimated at 12–15°C. The lowest temperature yielded larger and heavier adults (312 μg, dry weight) than did the highest temperature (225 μg). Fewer leafhoppers developed on the perennial than the annuals at 30°C and fewer on the annuals than the perennial at 18°C. Our results suggest that early in the season johnsongrass and perhaps other perennials are the superior developmental hosts for this leafhopper, whereas in midsummer when temperatures are highest, annuals are the better hosts.
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  • 2
    ISSN: 1432-0983
    Keywords: 2-oxoglutarate dehydrogenase ; Saccharomyces cerevisiae ; rad52-mediated chromosome loss
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Ogd1 mutants of Saccharomyces cerevisiae are deficient in mitochondrial 2-oxoglutarate dehydrogenase activity; they cannot grow on glycerol and produce an increased amount of organic acids during growth on glucose as substrate. Using gamma ray-induced rad52-mediated chromosome loss the ogd1 mutation can be assigned to chromosome IX. Tetrad analysis of crosses between ogd1 and other markers on chromosome IX revealed that the OGD1 gene maps on the left arm of this chromosome 1.9 cM from his5.
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  • 3
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Orotate phosphoribosyl transferase ; Nucleotide sequence-5-phosphoribosyl 1-pyrophosphate (5PRPP)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Orotate phosphoribosyl transferase (OPRTase) catalyses the transformation of orotate to OMP in the pyrimidine pathway. In the yeast Saccharomyces cerevisiae, the URA5 gene is known to encode this enzyme activity. In this paper we present the cloning and sequencing of a yeast gene, named URA10, encoding a second OPRTase enzyme. Comparison of the predicted amino acid sequences between URA5 and URA10 genes shows more than 75% similarity. These sequences have also been compared to those of Escherichia coli, Podospora anserina, Sordaria macrospora and Dictyostelium discoideum. Remarkable similarities in the primary structure of these proteins have been found. Gene disruption experiments revealed that URA10 gene expression is responsible for the leaky phenotype of a ura5 mutant. Assays of OPRTase activity in extracts from ura5 and ura10 mutants indicate that the URA10 product contributes only 20% of the total activity found in wild type cells.
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  • 4
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Mutants ; Farnesyl diphosphate synthetase ; Ergosterol
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Two yeast mutant strains auxotrophic for ergosterol and blocked in farnesyl diphosphate synthetase (EC 2.5.1.1) were isolated. Genetic analysis has shown that these mutant strains carry additional mutations in the ergosterol pathway besides erg20-1 and erg20-2 which affect FPP synthetase. The novel feature of these mutants is their ability to excrete prenyl alcohols (farnesol and geraniol). As geraniol is toxic for yeast cells, the above leaky mutations in FPP synthetase have to be associated with others in the sterol pathway, in order to slow down geraniol synthesis.
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  • 5
    ISSN: 1432-0983
    Keywords: Glucose oxidase ; Aspergillus ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We report the cloning of the Aspergillus niger glucose oxidase gene and its use to elevate glucose oxidase productivity in A. niger by increasing the gene dosage. In addition, the gene has been introduced into A. nidulans where it provides the novel capacity to produce glucose oxidase. A plasmid, in which DNA encoding the mature form of glucose oxidase was preceded by a Saccharomyces cerevisiae secretion signal, effected high-level production of extracellular glucose oxidase in this yeast.
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  • 6
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    Current genetics 18 (1990), S. 401-403 
    ISSN: 1432-0983
    Keywords: Baking yeast ; Saccharomyces cerevisiae ; Dough leavening ; Benomyl
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary To investigate the leavening ability of yeast in dough, chromosome loss was induced by benomyl treatment in YOY1037, a diploid between a baking strain and a laboratory strain, and its effect on the leavening ability was studied. When benomyl-treated cells were spread on plates with a dye indicator for ploidy, about 20% of the visible colonies were stained dark blue or dark purple; the rest stained pale blue, similar to the diploid YOY1037. Strains showing the MATα phenotype, and non-galactose fermenting strains, apparently having lost particular chromosomes, were observed only in those with darkcoloured colonies. Strains with dark-coloured colonies showed a wider range of leavening ability than did those with pale-coloured colonies.
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  • 7
    ISSN: 1432-0983
    Keywords: Xylitol dehydrogenase gene ; Pichia stipitis ; Saccharomyces cerevisiae ; Xylose utilization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A P. stipitis cDNA library in λgt11 was screened using antisera against P. stipitis xylose reductase and xylitol dehydrogenase, respectively. The resulting cDNA clones served as probes for screening a P. stipitis genomic library. The genomic XYL2 gene was isolated and the nucleotide sequence of the 1089 bp structural gene, and of adjacent non-coding regions, was determined. The XYL2 open-reading frame codes for a protein of 363 amino acids with a predicted molecular mass of 38.5 kDa. The XYL2 gene is actively expressed in S. cerevisiae transformants. S. cerevisiae cells transformed with a plasmid, pRD1, containing both the xylose reductase gene (XYL1) and the xylitol dehydrogenase gene (XYL2), were able to grow on xylose as a sole carbon source. In contrast to aerobic glucose metabolism, S. cerevisiae XYL1-XYL2 transformants utilize xylose almost entirely oxidatively.
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  • 8
    ISSN: 1432-0983
    Keywords: Mutagen hyper-resistance ; Nitrogen mustard ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A screening of haploid yeast strains for enhanced resistance to nitrogen mustard (HN2) yielded a recessive mutant allele, hnm1, that conferred hyper-resistance (HYR) to HN2. Diploids, homo- or heterozygous for the HNM1 locus, exhibit normal wild-type like resistance while homozygosity for hnm1 leads to the phenotype HYR to HN2. The hnm1 mutation could be found in yeast strains proficient or deficient in different DNA repair systems. In these mostly HN2-sensitive haploid repair-deficient mutants, hnm1 acted as a partial suppressor of HN2 sensitivity. All isolated recessive mutations conferring hyper-resistance belonged to a single complementations group. The HYR to HN2 phenotype was maximally expressed in growing cells and was associated with reduced mutability by HN2. HNM1 most probably controls uptake of HN2 which would be impaired in the hnm1 mutants.
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  • 9
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; G418 resistance ; Gene cartridges ; Heterologous Gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Coding sequence cartridges for aminoglycoside phosphotransferase (APT) were isolated from bacterial transposon Tn903. When incorporated into a heterologous gene construction utilising the PGK1 promoter and terminator, the heterologous APT gene provided a G418-resistance determinant that functioned efficiently as a dominant marker for yeast in both multiple- and single-copy. Transformant colonies on selective medium appeared rapidly, within 36–48 h, and growth rate of the transformed cells was normal. A simple and highly sensitive radiolabelling assay for APT enzyme activity was developed for use with crude cell protein extracts. Enzyme activity units were equated to the amount of APT protein present in the cells, and the APT protein was shown to be stable in yeast. Heterologous APT expression was 130-fold reduced compared with homologous PGK1. This resulted from an estimated two-fold decrease in mRNA level and a 65-fold decrease in translation efficiency. The latter was unaffected by AUG sequence context change, but corresponded with a high frequency of minor codons in the APT-coding sequence. APT can be used as a semi-quantitative reporter of gene expression, whose useful features are in vivo detection via the G418-resistance phenotype and powerful cell-free assay.
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  • 10
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Episomal plasmid ; Copy number control ; Plasmid maintenance ; Glycolytic enzyme levels
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary This study demonstrates how varying the promoter strength of an essential gene on a yeast 2μORI-STB YEp multicopy vector can influence vector copy levels. A phosphoglycerate kinase gene (PGK) on this plasmid was made essential for fermentative growth by transformation into a pgk - yeast strain. When in these PGK- transformants the requirement for PGK expression was the sole selective criterion for plasmid maintenance, PGK promoter activity was inversely related to vector copy levels. Plasmids with an efficiently-transcribed PGK gene were maintained at approximately one copy per cell, whereas those lacking the UAS that normally directs high basal PGK transcription levels were present at up to 10–15 copies. All cultures of these PGK+ transformants contained only a low proportion of pgk - cells. Since mitotic loss of the plasmid arrests growth through loss of a functional PGK allele, PGK confers high stability to the YEp vector in such a pgk - genetic background. In this system YEp vector levels are probably influenced by PGK transcription because high expression of PGK is needed in rapid fermentative growth. Remarkably, low plasmid PGK promoter activity caused PGK mRNA levels slightly higher than those found in yeast with normal PGK regulation. A higher plasmid copy number is therefore not the only factor counteracting the effects of low PGK transcription, and it is possible that PGK mRNA becomes more stable in response to inefficient PGK transcription.
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  • 11
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Sporulation ; Inessential genes ; Genome organization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The SPR6 gene of Saccharomyces cerevisiae encodes a moderately abundant RNA that is present at high levels only during sporulation. The gene contains a long open reading frame that could encode a hydrophilic protein approximately 21 kDa in size. This protein is probably produced by the yeast, because the lacZ gene of Escherichia coli is expressed during sporulation when fused to SPR6 in the expected reading frame. SPR6 is inessential for sporulation; mutants that lack SPR6 activity sporulate normally and produce viable ascospores. Nonetheless, the SPR6 gene encodes a function that is relevant to sporulating cells; the wild-type allele can enhance sporulation in strains that are defective for several SPR functions. SPR6 is located on chromosome V, 14.4 centimorgans centromere-distal to MET6.
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  • 12
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Nucleo-mitochondrial interactions ; Mitochondrial status ; Lycorine
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary In a previous paper we have shown that the alkaloid lycorine inhibits growth of rho +, mit - and rho -, strains of Saccharomyces cerevisiae, whereas strains devoid of mitochondrial DNA (rho o) are resistant to more than 200 μg/ml of the alkaloid. In this report we show that hypersuppressive petites are almost as resistant as rho o mutants, whereas isogenic rho - petites, which have retained tained longer segments of the genome, are sensitive to the drug.
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  • 13
    ISSN: 1432-0983
    Keywords: Schizosaccharomyces pombe ; Saccharomyces cerevisiae ; CaMV 35S promoter ; CaMV 35S terminator ; Heterologous expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Complementation of fission yeast mutants by plant genomic libraries could be a promising method for the isolation of novel plant genes. One important prerequisite is the functioning of plant promoters and terminators in Schizosaccharomyces pombe and Saccharomyces cerevisiae. Therefore, we studied the expression of the bacterial β-glucuronidase (GUS) reporter gene under the control of the Cauliflower Mosaic Virus (CaMV) 35S promoter and 35S terminator. We show here that S. pombe initiates transcription at exactly the same start site as was reported for tobacco. The 35S CaMV terminator is appropriately recognized leading to a polyadenylated mRNA of the same size as obtained in plant cells transformed with the same construct. Furthermore, the GUS-mRNA is translated into fully functional GUS protein, as determined by an enzymatic assay. Interestingly, expression of the 35S promoter in the budding yeast S. cerevisiae was found to be only moderate and about hundredfold lower than in S. pombe. To investigate whether different transcript stabilities are responsible for this enormous expression difference in the two yeasts, the 35S promoter was substituted by the ADH (alcohol dehydrogenase) promoter from fission yeast. In contrast to the differential expression pattern of the 35S promoter, the ADH promoter resulted in equally high expression rates in both fission and budding yeast, comparable to the 35S promoter in S. pombe. Since the copy number of the 35S-GUS constructs differs only by a factor of two in the two yeasts, it appears that differential recognition of the 35S promoter is responsible for the different transcription rates.
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  • 14
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; Mitochondria ; Intron-encoded proteins ; Recombination
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The respiratory competency of a yeast strain devoid of mitchondrial introns is quite normal. However, it may be asked whether intron-encoded proteins participate in metabolisms other than those of mitochondrial introns. Using strains without mitochondrial introns we have answered two questions. The first was: does the absence of intron-encoded proteins abolsh mitochondrial recombination? The second was: do mitochondrial introns and intron-encoded proteins play a part in mitochondrial DNA rearrangements induced by ethidium bromide (rho- production)? We have shown that the introns and intron-encoded proteins are not essential essential components of either phenomenon.
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  • 15
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    Current genetics 18 (1990), S. 23-27 
    ISSN: 1432-0983
    Keywords: Protein translocation ; Saccharomyces cerevisiae ; Peroxisomes ; Overexpression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Import of proteins into organelles usually requires a cis-acting targeting signal. Analysis of various hybrid proteins, consisting of mouse DHFR and parts of catalase A from Saccharomyces cerevisiae, revealed that fusion proteins containing the N-terminal 126 amino acids, or less, of catalase A remain in the cytosol whereas fusion proteins containing 140, or more, N-terminal amino acids of catalase A form large aggregates inside the cell. These protein bodies, which lack a surrounding membrane, copurified with peroxisomes on cell fractionation. The peroxisomal targeting signal of catalase A does not reside at the C-terminus or at the N-terminus.
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  • 16
    ISSN: 1573-0832
    Keywords: Nystatin ; amphotericin B ; amphotericin B methyl ester ; polyene antibiotics ; yeast ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Saccharomyces cerevisiae was cultured under anaerobiosis in semi-complete medium to which either palmitoleic or oleic acid was added. Cells were grown at 20 °C or 30 °C. The levels of total lipids, total sterols, and phospholipids were higher in cells grown at 20 °C than at 30 °C. The effects of nystatin (NYS), amphotericin B (AMB), and amphotericin B methyl ester (AME) were evaluated by determining cell viability and liberation of intracellular compounds. The loss of cell viability is higher in the first 30 minutes of incubation with the drugs and is the same regardless of the type of cells obtained. Low molecular weight compounds and ions such as K+ are liberated a few minutes after incubation with the drugs whereas proteins and substances absorbing at 260 nm are liberated later. Phosphate liberation comes after K+ and before compounds of higher molecular weights.
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  • 17
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    Nutrient cycling in agroecosystems 23 (1990), S. 73-80 
    ISSN: 1573-0867
    Keywords: Nigerian savanna ; maize ; potassium ; zinc
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract A three year field study was conducted at five locations in the Nigerian savanna to evaluate the response of early maturing maize variety to varying rates of K and Zn with a view to establishing the K and Zn requirements for maize production in this zone. Treatments consisted of 4 × 3 factorial combinations of 4 levels of K and 3 levels of Zn. Responses to K and Zn fertilization were sporadic and were obtained only in soils of the Southern Guinea savanna and in the soil formed on sedimentary sandstone. There seem to be no problem at present in soils of the Northern Guinea and Sudan savannas where leaching is less intense. It is inferred from this study that K and Zn deficiences are incipient in the high rainfall soils and in the sandstone derived soils. For these soils, 50 kg K/ha and 2–5 kg Zn/ha is suggested as adequate for an early maturing maize crop. Soil data showed that K and Zn responses can be expected when available K and Zn levels fall below 0.1 meq/100 g and 2 ppm respectively.
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  • 18
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    Nutrient cycling in agroecosystems 26 (1990), S. 249-252 
    ISSN: 1573-0867
    Keywords: Long-term manure trial ; residual effect ; model test ; nitrogen availability ; maize ; Italian ryegrass
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Sluijsmans and Kolenbrander developed a simple model to describe the availability of animal manure, assuming a readily available, an easily decomposable and a slowly decomposable N fraction. We tested this model on data from an experiment in which farmyard manure had been applied for eleven successive years to silage maize [Zea mays L.] grown on a light sandy soil. The residual effects of this FYM were then measured by growing Italian ryegrass [Lolium multiflorum Lamk.] in the 12th year. The measured uptake of N by the grass of the FYM residues was then compared with the computed values. The measured amounts of N taken up agreed fairly well with the calculated amounts for applications of 50 and 100 t FYM per ha per year. If the rates of manure application are adjusted to crop requirement, the model shows that the potential, long-term release of N from the residual N fraction of FYM will not exceed 20 kg N per ha. For cattle slurry with a smaller residual fraction, the release will be at most 10% of the total annual N application.
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  • 19
    ISSN: 1572-9699
    Keywords: 2-Deoxy-D-glucose transport ; polyphosphate ; Saccharomyces cerevisiae ; sugar phosphorylation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The role of polyphosphate in 2-deoxy-D-glucose transport was studied in yeast cells, pulse-labeled with [32P]orthophosphate, by comparing the concentrations and specific activities of polyphosphate, orthophosphate and 2-dGlc-phosphate. When 2-dGlc transport was measured under aerobic conditions, it appeared that polyphosphate replenished the orthophosphate pool, indicating that polyphosphate has, at least mainly, an indirect role in sugar phosphorylation. Also in cells with a reduced respiratory capacity, due to a treatment with antimycin A, no direct role for polyphosphate in 2-dGlc transport could be detected. Under these conditions, only a very limited breakdown of polyphosphate occurred, probably because of the small decrease in the orthophosphate concentration.
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  • 20
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    Cellular and molecular life sciences 46 (1990), S. 193-200 
    ISSN: 1420-9071
    Keywords: Saccharomyces cerevisiae ; protein toxin ; yeast toxin precursor ; protease processing ; lectin ; (1→6)-β-D-glucan ; receptor ; resistant mutants ; spheroplasts ; ion-permeable channels ; site-directed mutagenesis ; toxin functional domains
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The K1 killer toxin ofSaccharomyces cerevisiae is a secreted, virally-coded protein lethal to sensitive yeasts. Killer yeasts are immune to the toxin they produce. This killer system has been extensively examined from genetic and molecular perspectives. Here we review the biology of killer yeasts, and examine the synthesis and action of the protein toxin and the immunity component. We summarise the structure of the toxin precursor gene and its protein products, outline the proteolytic processing of the toxin subunits from the precursor, and their passage through the yeast secretory pathway. We then discuss the mode of action of the toxin, its lectin-like interaction with a cell wall glucan, and its probable role in forming channels in the yeast plasma membrane. In addition we describe models of how a toxin precursor species functions as the immunity component, probably by interfering with channel formation. We conclude with a review of the functional domains of the toxin structural gene as determined by site-directed mutagenesis. This work has identified regions associated with glucan binding, toxin activity, and immunity.
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  • 21
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    Archives of microbiology 154 (1990), S. 267-273 
    ISSN: 1432-072X
    Keywords: Yeast ; Saccharomyces cerevisiae ; (R)-2,3-Butanediol dehydrogenase ; Stereospecificity ; Gas chromatographic analysis of enantiomers
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A NAD-dependent (R)-2,3-butanediol dehydrogenase (EC 1.1.1.4), selectively catalyzing the oxidation at the (R)-center of 2,3-butanediol irrespective of the absolute configuration of the other carbinol center, was isolated from cell extracts of the yeast Saccharomyces cerevisiae. Purification was achieved by means of streptomycin sulfate treatment, Sephadex G-25 filtration, DEAE-Sepharose CL-6B chromatography, affinity chromatography on Matrex Gel Blue A and Superose 6 prep grade chromatography leading to a 70-fold enrichment of the specific activity with 44% yield. Analysis of chiral products was carried out by gas chromatographic methods via pre-chromatographic derivatization and resolution of corresponding diasteromeric derivatives. The enzyme was capable to reduce irreversibly diacetyl (2,3-butanediol) to (R)-acetoin (3-hydroxy-2-butanone) and in a subsequent reaction reversibly to (R,R)-2,3-butanediol using NADH as coenzyme. 1-Hydroxy-2-ketones and C5-acyloins were also accepted as substrates, whereas the enzyme was inactive towards the reduction of acetone and dihydroxyacetone. The relative molecular mass (M r) of the enzyme was estimated as 140 000 by means of gel filtration. On SDS-polyacrylamide gel the protein decomposed into 4 (identical) subunits of M r 35 000. Optimum pH was 6.7 for the reduction of acetoin to 2,3-butanediol and 7.2 for the reverse reaction.
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  • 22
    ISSN: 1432-072X
    Keywords: cAMP ; Cat mutants ; Glucose repression ; Glucose-induced ; Intracellular pH ; Ras ; Saccharomyces cerevisiae ; Signal transduction ; Trehalase ; Yeast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Addition of glucose to derepressed cells of the yeast Saccharomyces cerevisiae induces a transient, specific cAMP signal. Intracellular acidification in these cells, as caused by addition of protonophores like 2,4-dinitrophenol (DNP) causes a large, lasting increase in the cAMP level. The effect of glucose and DNP was investigated in glucose-repressed wild type cells and in cells of two mutants which are deficient in derepression of glucose-repressible proteins, cat1 and cat3. Addition of glucose to cells of the cat3 mutant caused a transient increase in the cAMP level whereas cells of the cat1 mutant and in most cases also repressed wild type cells did not respond to glucose addition with a cAMP increase. The glucose-induced cAMP increase in cat3 cells and the cAMP increase occasionally present in repressed wild type cells however could be prevented completely by addition of a very low level of glucose in advance. In derepressed wild type cells this does not prevent the specific glucose-induced cAMP signal at all. These results indicate that repressed cells do not show a true glucose-induced cAMP signal. When DNP was added to glucose-repressed wild type cells or to cells of the cat1 and cat3 mutants no cAMP increase was observed. Addition of a very low level of glucose before the DNP restored the cAMP increase which points to lack of ATP as the cause for the absence of the DNP effect. These data show that intracellular acidification is able to enhance the cAMP level in repressed cells. The glucose-induced artefactual increase occasionally observed in repressed cells is probably caused by the fact that their low intracellular pH is only restored after the ATP level has increased to such an extent that it is no longer limiting for cAMP synthesis. It is unclear why the artefactual increases are not always observed. Measurement of glucose- and DNP-induced activation of trehalase confirmed the physiological validity of the changes observed in the cAMP level. Our results are consistent with the idea that the glucose-induced signaling pathway contains a glucose-repressible protein and that the protein is located before the point where intracellular acidification triggers activation of the pathway.
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  • 23
    ISSN: 1432-072X
    Keywords: Saccharomyces cerevisiae ; Catalase A ; Catalase T ; β-Oxidation ; Microbodies ; H2O2-Metabolism
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The parental strain (A+T+) of Saccharomyces cerevisiae and mutants, deficient in catalase T (A+T−), catalase A (A−T+) or both catalases (A−T−), grew on ethanol and oleic acid with comparable doubling times. Specific activities of catalase were low in glucose- and ethanol-grown cells. In the two oleic acid-grown A+-strains (A+T+ and A+T−) high catalase activities were found; catalase activity invariably remained low in the A−T+ strain and was never detected in the A−T− strain. The levels of β-oxidation enzymes in oleic acid-grown cells of the parental and all mutant strains were not significantly different. However, cytochrome C peroxidase activity had increased 8-fold in oleic acid grown A− strains (A−T+ and A−T−) compared to parental strain cells. The degree of peroxisomal proliferation was comparable among the different strains. Catalase A was shown to be located in peroxisomes. Catalase T is most probably cytosolic in nature and/or present in the periplasmic space.
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  • 24
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    Archives of microbiology 153 (1990), S. 384-391 
    ISSN: 1432-072X
    Keywords: Saccharomyces cerevisiae ; Ethanol ; Acetic acid ; Cytoplasmic pH ; 31P-NMR ; 13C-NMR
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    Topics: Biology
    Notes: Abstract Cell suspensions of a respiratory deficient mutant of Saccharomyces cerevisiae were monitored by in vivo 31P and 13C Nuclear Magnetic Resonance in order to evaluate the effect of ethanol in intracellular pH and metabolism. In the absence of an added energy source, ethanol caused acidification of the cytoplasm, as indicated by the shift to higher field of the resonance assigned to the cytoplasmic orthophosphate. Under the experimental conditions used this acidification was not a consequence of an increase in the passive influx of H+. With cells energized with glucose, a lower value for the cytoplasmic pH was also observed, when ethanol was added. Furthermore, lower levels of phosphomonoesters were detected in the presence of ethanol, indicating that an early event in glycolysis is an important target of the ethanol action. Acetic acid was identified as responsible for the acidification of the cytoplasm, in experiments where [13C]ethanol was added and formation of labeled acetic acid was detected. The intracellular and the extracellular concentrations of acetic acid were respectively, 30 mM and 2 mM when 0.5% (120 mM) [13C]ethanol was added.
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  • 25
    ISSN: 1432-072X
    Keywords: Regulatory mutants ; Meiotic mapping ; Transcriptional regulation ; MAL genes ; Saccharomyces cerevisiae
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    Notes: Abstract The MAL1 locus of Saccharomyces cerevisiae comprises three genes necessary for maltose utilization: a regulatory (MALR), a maltose transport (MALT) and a maltase gene (MALS). A fine structure genetic map of the MAL1R gene was constructed and the order of mutations was confirmed by plasmid-mediated chromosomal recombination. The mutations cluster non-randomly within the 5′ half of the gene, where the putative DNA binding domain of the encoded protein is located. Only mutations mal1 R-22 and MAL1R-72 map in the 3′ terminal half of the gene; these mutations cause a different pattern of transcriptional regulation of plasmid-borne MAL6T genes. Experiments supporting a direct involvement of the MALR-encoded protein in carbon catabolite repression of MAL gene expression are reported.
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  • 26
    ISSN: 1573-4927
    Keywords: mutator ; transposable element ; alcohol dehydrogenase ; maize ; gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract A secondary mutant, derived from an allele of maize alcohol dehydrogenase 1 (Adh1) carrying a Mutator transposable element (Mu1) in its first intron, was reported to exhibit a threefold decrease in ADH enzymatic activity and steady-state RNA levels compared to the original mutant. The original mutant,Adh1-S3034 (abbreviatedS3034), was previously characterized at the molecular level. The derivative, abbreviatedS3034b, has now been cloned; at the DNA sequence level the insertion and surroundingAdh1 sequences are indistinguishable fromS3034. Furthermore, in our lines there is no difference in relative ADH activities between products of the two putative alleles. A comparison of gene expression in heterozygotes obtained by crossing to different tester lines reveals a correlation between the measured decrease in levels of ADH polypeptide produced by the mutant allele and the background in which it is measured; this effect is distinct from any background-related variation in the expression of the progenitor allele. It does not appear to be attributable to alternative patterns of DNA modification. It appears to reflect a background-associated difference in the level of normalAdh1-RNA produced. Thus the previously reported distinction betweenS3034 andS3034b may be due to differences in the extent to which the mutant allele and a given genetic background interact to produce functionalAdh1-RNA.
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  • 27
    ISSN: 1573-4919
    Keywords: bovine heart fatty acid-binding protein ; H-FABPc ; heterologous gene expression ; Saccharomyces cerevisiae ; GALIO promoter
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Summary The unicellular eukaryotic microorganism, Saccharomyces cerevisiae, transformed with a plasmid containing a cDNA fragment encoding bovine heart fatty acid-binding protein (H-FABP) under the control of the inducible yeast GAL10 promoter, expressed FABP during growth on galactose. The maximum level of immunoreactive FABP, identical in size to native protein as judged from SDS-polyacrylamide gel electrophoresis, was reached after approximately 16 hours of induction. Analysis of particulate and soluble subcellular fractions showed that FABP was exclusively associated with the cytosol. FABP expressed in yeast cells was functional as was demonstrated by its capacity to bind 14C-oleic acid in an in vitro assay. Growth of the transformants on galactose as the carbon source was significantly retarded at 37°C. Whereas the fatty acid pattern of total lipids was not altered in transformed cells, desaturation of exogenously added 14C-palmitic acid was significantly reduced both at 30 and 37°C. The lowest percentage of radioactively labeled unsaturated fatty acids was found in the phospholipid fraction.
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  • 28
    ISSN: 1573-1561
    Keywords: Leafhopper ; Dalbulus maidis ; Homoptera ; Cicadellidae ; hostfinding ; maize ; visual ; olfactory ; synergism ; pest
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Virtually nothing is known about the role plant volatiles play in host-finding by Homoptera in the Suborder Auchenorrhyncha. In laboratory bioassays, we examined the influence of plant volatiles on orientation and postcontact behaviors of the leafhopper,Dalbulus maidis, and determined the relationship between visual and olfactory stimuli during host-finding. When compared to the number of contacts made with reflected green light in the presence of a hexane control,D. maidis made more contacts when exposed to volatile extracts from its preferred host, maize; a similar number of contacts when exposed to volatiles from a marginal host, gamagrass; and fewer contacts when exposed to volatiles from a nonhost, sorghum. There was no difference between males and females in the number of contacts made with green light when exposed to maize volatiles compared to hexane alone. More contacts were made with green light than with white light of similar intensity, both in the presence and in the absence of olfactory stimuli; however, maize volatiles acted as a Synergist by increasing the number of contacts leafhoppers made with green light. After contacting the green light, exposure of maize volatiles significantly increased, relative to hexane, the amount of stationary time, but did not influence the amount of time spent moving, the distance traveled, or the speed while moving when within the boundaries of the green light. This study provides the first evidence for an interaction between visual and olfactory stimuli during host-finding for a leafhopper and also for olfactory mediation of postcontact behaviors not associated with feeding.
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  • 29
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    Plant molecular biology 14 (1990), S. 163-171 
    ISSN: 1573-5028
    Keywords: rice ; actin ; cDNA sequence ; transcript mapping ; gene structure
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    Topics: Biology
    Notes: Abstract We have isolated and sequenced a full-length cDNA clone containing information for the rice actin gene RAc1. Transcript terminus mapping and sequence alignment between the RAc1 cDNA clone and a previously isolated RAc1 genomic clone were used to determine the structure of the RAc1 gene. This allowed us to make the first complete structural characterization of a plant actin gene. The analysis revealed the presence of a 5′-noncoding exon, separated by an intron, from the first translated exon of the RAc1 gene. This is one of the few reported cases of a plant gene containing such a 5′-noncoding exon. Sequence comparison between the previously isolated plant actin genes suggests that such an exon may be a common feature of plant actin gene structure. The present study also confirms that the rice actin gene family is composed of at least eight unique members.
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  • 30
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    Plant molecular biology 14 (1990), S. 333-347 
    ISSN: 1573-5028
    Keywords: intergenic spacer ; maize ; methylation ; rDNA ; teosinte
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The restriction endonucleases Hpa II and Msp I were used to examine cytosine methylation in the ribosomal RNA genes (rDNA) of inbred lines of maize and species of teosinte. In all of the rDNAs examined, Msp I (not sensitive to mCpG) digestion yielded a distribution of lower molecular weight fragments indicative of multiple recognition sites. The majority of the rDNA arrays in an individual were inaccessible to Hpa II (sensitive to mCpG) cleavage, but a significant fraction (10–25%) was cleaved at least once by Hpa II into repeat unit length fragments (9.1 kbp). In some maize inbred lines, one or two additional fragment populations (less than 9.1 kbp in length) were also produced by Hpa II digestion. All of the unmethylated Hpa II sites mapped to the intergenic spacer (IGS), and the major unmethylated site was located approximately 800 bp 5′ to the start of the 18S RNA coding sequence. An Eco RI polymorphism, present in the 26S gene of certain inbred lines and hybrids, was utilized to investigate the organization of unmethylated repeat units in the rDNA array. In double digest experiments with Hpa II/Eco RI, the fragments from repeat units with two Eco RI sites were sensitive to Hpa II digestion, whereas, the fragments from repeat units with a single Eco RI site were almost completely resistant to Hpa II digestion. Similar digestion patterns were also observed in Eco RII (sensitive to mCNG)/Eco RI digests. These results suggest that unmethylated and Eco RI polymorphic sites occur in the same repeat units.
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  • 31
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    Plant molecular biology 15 (1990), S. 399-406 
    ISSN: 1573-5028
    Keywords: light-inducible ; rice ; shoot-specific
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    Topics: Biology
    Notes: Abstract By differential screening of a cDNA library of two-week-old rice seedlings cDNA clones were obtained, corresponding to shoot-specific mRNAs. By sequence analysis two of these clones were found to be rbcS cDNA clones. The mRNA corresponding to a third cDNA clone (COS5) displayed an expression pattern similar to the expression pattern of rbcS genes. The mRNA (800 bases) was light-inducible and encoded by a single-copy gene. The genomic clone (GOS5) was isolated and the intron/exon structure was determined by comparing the nucleotide sequence of the mRNA and the genomic clone. The gene contains two introns. Transcription start sites were determined by S1-nuclease mapping and primer extension. The start site obtained by both methods is located 87 bp upstream of the translation start site and 23 bp downstream of TATA box-like sequence. In the 5′ non-coding region motifs can be found that are homologous to sequences in promoters that are light-or UV-inducible or confer leaf-specific expression. The open reading frame present in GOS5 codes for a protein (15 kDa) that contains a putative chloroplast transit peptide and does not show any significant homology to protein sequences in the NBRF protein database.
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  • 32
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    Plant molecular biology 15 (1990), S. 783-785 
    ISSN: 1573-5028
    Keywords: sorghum ; maize ; glycine-rich proteins ; RNA-binding
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  • 33
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    Plant molecular biology 14 (1990), S. 1-15 
    ISSN: 1573-5028
    Keywords: cDNA ; differential screening ; genomic cloning ; maize ; tandem genes ; α-tubulin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The identification of a cDNA (MR19) corresponding to a maize α-tubulin and homologous genomic clones (MG19/6 and MG19/14) is described. The cDNA has been isolated by differential screening of a cDNA maize root library. We have found two α-tubulin genes in a tandem arrangement in the genomic clones, separated by approximately 1.5 kbp. One of the genes (gene I) contains an identical nucleotide sequence which corresponds to the cDNA clone. The two deduced proteins from DNA sequences are very similar (only two conservative replacements in 451 amino acids) and they share a high homology as compared with the published α-tubulin sequences from other systems and in particular with the Arabidopsis thaliana and Chlamydomonas reinhardtii sequences reported. The structure of both genes is also very similar; it includes two introns, of 1.7 kbp and 0.8 kbp respectively, in each gene and only one intron placed at a homologous position in relation to Arabidopsis thaliana genes. By using specific 3′ probes it appears that both genes are preferentially expressed in the radicular system of the plant. The α-tubulin gene family of Zea mays seems to be represented by at least 3 or 4 members.
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  • 34
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    Plant molecular biology 14 (1990), S. 29-39 
    ISSN: 1573-5028
    Keywords: abscisic acid ; gene locus ; gene sequence ; osmoregulation ; rice
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    Topics: Biology
    Notes: Abstract We have cloned and sequenced the four members of a rice rab (responsive to abscisic acid) gene family that are tandemly arrayed in a locus approximately 30 kbp in length. Each of the genes contains a single, small intron. They are all transcriptionally active and encode proteins of M r 15500–16800 with two highly conserved domains. Northern analysis with gene-specific probes showed slightly different patterns of expression for the four genes in rice plant tissues and in response to osmotic stress. Comparison of the promoter regions revealed a conserved GC-rich sequence (CGG/CCGCGCT) with some homology to the SP1 binding site (Briggs et al., 1986). Another conserved sequence (PuTACGTGGCPu), whose core is found in the promoter regions of ABA-responsive contton genes, is reminiscent of the cAMP responsive element (Deutsch et al., 1988).
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  • 35
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    Plant molecular biology 14 (1990), S. 621-624 
    ISSN: 1573-5028
    Keywords: rice ; actin ; genomic DNA sequences ; sequence identity
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  • 36
    ISSN: 1573-5028
    Keywords: maize ; Coix ; zein ; coixin ; phylogenetic relationship
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    Topics: Biology
    Notes: Abstract Zeins from Zea mays L cv. Maya and coixins from Coix lacryma-jobi L. cv. Adlay were fractionated to obtain α-, β-, and γ-zein and α-, β-, and γ-coixin. The α-coixins were composed of 4 polypeptide classes of 27 kDa (C1), 25 kDa (C2), 17 kDa (C4) and 15 kDa (C5) with solubility properties very similar to those of the 22 kDa and 19 kDa α-zeins. Like the α-zeins, the C1 and C2 α-coixins corresponded to 80% of total Coix prolamins. The fraction corresponding to γ-coixin contained only one protein band of 22 kDa (C3). This coixin fraction has solubility properties similar to those of γ-zein and represents 15% of the total coixin. The β-zein fraction was composed of a major 17 kDa protein band, while the β-coixin fraction consisted of a mixture of α- and γ-coixins. Polyclonal antibodies raised against C1 recognized C1 and C2 and cross-reacted strongly with the 22 kDa α-zein, as did C4 and C5 antisera. The antiserum against γ-coixin showed strong cross-reaction with γ-zein. The homology between coixins and zeins was further investigated by using Southern hybridization analyses. The genomic DNA of maize and Coix were digested with several restriction enzymes and probed with cDNA clones representing 19 and 22 kDa α-zeins as well as the 28 and 16 kDa γ-zeins. The Coix genome showed complex cross-hybridization sequences with the 22 kDa α-zein cDNA, while no cross-hybridization was observed with the 19 kDa cDNA clone. The cDNA clone representing the 28 kDa γ-zein cross-hybridized with only one band of Coix genomic DNA, in contrast to the three bands observed in maize. This same Coix sequence also cross-hybridized with the cDNA clone representing the 16 kDa γ-zein. The relevance of these findings are discussed in the context of the origin of zein and coixin genes.
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  • 37
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    Plant molecular biology 15 (1990), S. 257-268 
    ISSN: 1573-5028
    Keywords: actin genes ; differential expression ; gene evolution ; gene family ; rice
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Four rice (Oryza sativa) genomic actin genes have been characterized. The rice actin genes show a conservation of intron number and position that is characteristic of plant actins. Sequence comparisons revealed that the plant actins generally have a high degree of inter- and intraspecific sequence heterogeneity. However, one rice actin gene has a much higher degree of nucleotide sequence identity to a previously isolated actin sequence from Arabidopsis thaliana than to any other plant actin gene. This leads us to suggest that the two sequences may represent functionally homologous genes which arose from an ancient actin gene lineage that was separated by the divergence of the dicot and monocot plants. Genomic DNA blot analysis showed that the rice actin gene family contains at least eight unique members. RNA hybridization analysis revealed that individual rice actin genes can display different patterns of transcript accumulation. The observed differences in sequence and transcript accumulation patterns suggest that the individual rice actin genes may differ in their transcriptional regulation and/or cellular function.
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  • 38
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    Plant molecular biology 15 (1990), S. 905-912 
    ISSN: 1573-5028
    Keywords: rice ; developmental regulation ; embryo-specific
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    Topics: Biology
    Notes: Abstract We have analyzed in transgenic tobacco the expression of a chimeric gene containing 5′ sequences of the rice rab-16B gene fused to the β-glucuronidase (GUS) reporter gene. This construct, a translational fusion (−482 to +184) including 14 amino acids of the RAB-16B protein, is expressed only in zygotic and pollen-derived embryos. In zygotic embryos, GUS activity begins to accumulate 10 days after flowering (daf), and increases until seed maturation at 25 daf. Immunological measurements of endogenous abscisic acid (ABA) accumulation in these seeds showed a close parallel between hormone levels and GUS activity. However, GUS activity could not be reproducibly induced by treatment of immature embryos with ABA (10 μM). Neither GUS activity nor GUS mRNA could be detected in leaves of transgenic tobacco even after ABA treatment. In contrast, GUS activity could be induced to high levels in pollen-derived embryos by treatment with ABA. Our results show that 482 bp of 5′ sequences of the rice rab-16B promoter can confer in transgenic tobacco developmentally regulated expression in embryos but not ABA-responsive expression in vegetative tissues.
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  • 39
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    Plant molecular biology 15 (1990), S. 933-935 
    ISSN: 1573-5028
    Keywords: rice ; rRNA gene ; intergenic spacer
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  • 40
    ISSN: 1573-5028
    Keywords: maize ; ABA-induced gene ; protein phosphorylation
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    Topics: Biology
    Notes: Abstract The ABA-induced MA12 cDNA from maize, which encodes a set of highly phosphorylated embryo proteins, was used to isolate the corresponding genomic clone. This gene, called RAB-17 (responsive to ABA), encodes a basic, glycine-rich protein (mol. wt. 17 164) containing a cluster of 8 serine residues, seven of them contiguous. It is a homologue of the rice RAB-21 gene (Mundy J, Chua NH, EMBO J 7; 2279–2286, 1988). Phosphoamino acid analysis of the isolated protein indicates that only the serine residues are phosphorylated and a putative casein-type kinase phosphorylatable sequence was identified in the protein. The pattern of expression and in vivo phosphorylation of the RAB-17 protein was studied during maize embryo germination and in calli of both meristematic or embryonic origin. ABA treatment induced the synthesis of RAB-17 mRNA and protein in calli, however, the RAB-17 proteins were found to be highly phosphorylated only in embryos.
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  • 41
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    Plant molecular biology 15 (1990), S. 913-920 
    ISSN: 1573-5028
    Keywords: Adh1 ; chloramphenicol acetyl transferase ; enhancer ; intron ; maize ; transient expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Chimeric genes containing the coding sequence for bacterial chloramphenicol acetyl transferase (CAT) have been introduced by electroporation into maize protoplasts (Black Mexican Sweet) and transient expression monitored by enzyme assays. Levels of CAT expression were enhanced 12-fold and 20-fold respectively by the inclusion of maize alcohol dehydrogenase-1 introns 2 and 6 in the chimeric construct. This enhancement was seen when the intron was placed within the 5′ translated region but not when it was located upstream of the promoter or within the 3′ untranslated region. Deletion of exon sequences adjacent to intron 2 abolished its ability to mediate enhancement of CAT gene expression. Northern analysis of protoplasts electroporated with intron constructs revealed elevated levels of CAT mRNA. However, this elevation was insufficient to account for the increased enzyme activity. One explanation of these results is that splicing affects both the quantity of mRNA.
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  • 42
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    The journal of membrane biology 116 (1990), S. 93-105 
    ISSN: 1432-1424
    Keywords: clathrin ; genetics ; Saccharomyces cerevisiae ; exocytosis ; endocytosis ; prohormone maturation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
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  • 43
    ISSN: 1432-0878
    Keywords: Immunocytochemistry ; Met-enkephalin-Arg6-Phe7 ; Met-enkephalin-Arg6-Gly7-Leu8 ; Neurosecretory cells of insects ; Neuropeptides ; Co-existence of peptides ; Blowfly,Calliphora vomitoria (Insecta)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Neuronal pathways in the retrocerebral complex and thoracico-abdominal ganglionic mass of the blowflyCalliphora vomitoria have been identified immunocytochemically with antisera against the extended-enkephalins, Met-enkephalin-Arg6-Phe7 (Met-7) and Met-enkephalin-Arg6-Gly7-Leu8 (Met-8). Neurons of the hypocerebral ganglion, immunoreactive to Met-8, have axons in the crop duct nerve and terminals in muscles of the crop and its duct. Certain neurons of the hypocerebral ganglion are also immunoreactive to Met-7, and axons from these cells innervate the heart. Met-8 immunoreactive nerve terminals invest the cells of the corpus allatum. The source of this material is believed to ve a single pair of lateral neurosecretory cells in the brain. There is no Met-7 immunoreactive material in the corpus allatum. In the corpus cardiacum neither Met-7 nor Met-8 immunoreactivity is present in the cells. However, in the neuropil of the gland certain fibres, with their origins elsewhere, do contain Met-8 immunoreactivity. The most prominent neurons in the thoracic ganglion are the Met-7 immunoreactive ventral thoracic neurosecretory cells, axons from which project to neurohaemal areas in the dorsal neural sheath and also, via the ventral connective, to the brain. Co-localisation studies show that the perikarya of these cells are immunoreactive to antisera raised against several vertebrate-type peptides, such as Met-7, gastrin/cholecystokinin and pancreatic polypeptide. However, their axons and terminals show varying amounts of the peptides, suggesting differential transport and utilisation. Only a few cells in the thoracic ganglion are immunoreactive to Met-8 antisera. These lie close to the nerve bundles suppling the legs. In the abdominal ganglion, Met-8 immunoreactive neurons project to the muscles of the hindgut. This study suggests that the extended enkephalin-like peptides ofCalliphora may have a variety of different roles: as neurotransmitter or neuromodulator substances; in the direct innervation of effector organs; and as neurohormones.
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  • 44
    ISSN: 1432-0878
    Keywords: Insulin-like peptide ; Immunocytochemistry ; Immunochemical characterization ; Brain ; Neuroendocrine structures ; Leucophaea maderae (Insecta)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Immunocytochemical tests with eight monoclonal antibodies against either bovine or human insulin and seven polyclonal antibodies against bovine insulin were carried out to determine the presence of insulin-like neuropeptides in the brain and affiliated neuroendocrine structures of the insect Leucophaea maderae. Reaction products identified in the brain, subesophageal ganglion, and corpus cardiacum-corpus allatum complex indicate the presence of materials resembling mammalian insulins in its antigenic properties. The immunostaining observed with monoclonal antibodies appears to indicate the occurrence of an insulin-related peptide that shows sequential similarities with parts of both the A- and B-chains of mammalian insulin molecules. These suppositions are supported by the results of dot-blot and two-site time-resolved immunofluorometric assay (TR-IFMA) screenings of fractions of Leucophaea tissue extracts obtained by chromatography. The polyclonal antibodies yielded reaction products in some of the same areas and in additional parts of the neuroendocrine system not visualized by the monoclonal antibodies. Immunoreaction was observed in the following areas: the pars intercerebralis of the protocerebrum, the nervi corporis cardiaci I transporting insulin-like material to the corpus cardiacum, the dorsolateral protocerebral area and the optic lobes, the deutocerebrum, the tritocerebrum, and the subesophageal ganglion. In addition, smaller cell bodies with immunoreactive deposits occur at the border between proto- and deutocerebrum, and in the central area of the protocerebrum. The distribution of reactive material in the corpus cardiacum-corpus allatum complex after use of both groups of antibodies was the same. The fact that polyclonal and monoclonal antibodies yielded reaction products in different cells of the brain suggests either that the two groups of antibodies recognize different epitopes of the same molecule, or that they reveal two different types of immunoreactive molecules related to mammalian insulins. Together with the biochemical data reported by Nagasawa and coworkers (PNAS 83, 1986) the present immunocytochemical analysis has established a closer relationship between mammalian and insect “insulins” than was previously known.
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  • 45
    ISSN: 1432-0878
    Keywords: Serotonin ; Urotensins ; Somatostatin ; Immunocytochemistry ; Caudal neurosecretory system ; Reissner's fiber (subcommissural organ) ; Salmon,Oncorhynchus kisutch (Teleostei)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The caudal spinal cord of the coho salmon was investigated by means of immunocytochemistry using antisera against serotonin, urotensin I, urotensin II, somatostatin and a urea-extract of bovine Reissner's fiber (AFRU). Populations of serotonin-immunoreactive (IR) neurons were found rostral and dorsal to the urophysis in close spatial association with caudal secretory neurons. Thick, smooth serotonin-IR processes extended toward the external surface of the spinal cord where they displayed conspicuous terminal dilatations. Thin, beaded serotonin-IR fibers appeared to innervate populations of caudal secretory and somatostatin-IR cerebrospinal fluid-contacting neurons. Most caudal neurosecretory cells displayed both urotensin I and urotensin II immunoreactivities; only a minority reacted exclusively with either urotensin I or urotensin II antisera. Urotensin II-IR and somatostatin-IR cerebrospinal fluid (CSF)-contacting neurons were found as an integral component of the central canal wall in the caudal spinal cord and filum terminale; their dendritic processes appeared to contact Reissner's fiber, which displayed a weak AFRU-immunoreactivity while inside the central canal, but became strongly reactive in the interior of the terminal ventricle as it formed the massa caudalis. The distribution of serotoninergic processes points to a regulatory role in the function of caudal secretory and CSF-contacting neurons and to a putative serotonin release into the subarachnoid space and/or meningeal vasculature. It is also suggested that the CSF-contacting neurons of the central canal may participate in a feedback mechanism controlling the secretory activity of the subcommissural organ.
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  • 46
    ISSN: 1432-0878
    Keywords: Adenohypophysis ; Pars tuberalis ; Immunocytochemistry ; Thyroid-stimulating hormone (TSH) ; Propylthiouracil (PTU) ; Thyroxine (T4) ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The pars tuberalis (pt) of the adenohypophysis is unique in its close spatial relationship to the neurohemal contact area of the median eminence. The morphology of pt-specific secretory cells does not resemble cell types of the pars distalis (pd); the functional role of these cells within the endocrine system is still unknown. One group of young mature female Wistar rats received propylthiouracil (PTU), a second group thyroxine (T4) (10 mg/l each in drinking water) from about 3 weeks prior to the expected pregnancy and throughout the experiment. On gestation day 20, the fetuses were obtained by laparatomy. Serial sections from the rostral portion of the pt and from the pd were immunostained using the peroxidase-antiperoxidase method. TSH concentrations were determined by RIA in serum and pituitaries; T4 was measured in serum. An antiserum against rat (r) TSH revealed a moderate positive reaction of nearly all cells of the pt in the control group. In both experimental groups the pt-specific cells showed weak or no immunoreactivity. Sections of all groups were negative with anti(r)-LH,-GH,-PRL. In contrast to controls, only a few immature TSH-cells could be found in sections of the pd in the T4-group, while concentrations of TSH in blood and hypophysis were very low. TSH-cells in the PTU-group were enlarged and less intensely stained. TSH-concentrations were decreased in the hypophysis, blood levels were elevated. All sections of the pd-specific cell populations showed positive immunoreactions with anti(r)-LH,-GH,-PRL. The present results suggest that pt-specific secretory cells of the fetal rat possess TSH immunoreactivity but do not resemble the thyrotropes of the pd. Marked differences in immunoreactivity displayed by the experimental groups indicate that pt-specific cells respond to changes in the fetal thyroid status and are a component of the thyroid-regulating system in addition to the thyrotropes of the pd. This novel aspect of pt function is discussed in connection with recent results concerning melatonin receptors found in the pt and the inhibitory influence of the pineal gland exerted on the thyroid gland.
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    Cell & tissue research 260 (1990), S. 479-484 
    ISSN: 1432-0878
    Keywords: Histamine ; Immunocytochemistry ; Nervous system ; Excretory system ; Flatworms (Platyhelminthes) ; Diphyllobothrium dendriticum (Cestoda) ; Microstomum lineare (Turbellaria)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Specific antibodies against histamine were used to demonstrate the occurrence and cellular distribution of histamine-like immunoreactivity in three species of flatworms (phylum Platyhelminthes). In the parasitic cestode Diphyllobothrium dendriticum, histamine-reactivity was found in neurons of the main nerve cords, and in cells lining the central and peripheral excretory ducts. In the free-living microturbellarian Microstomum lineare and in the planarian Polycelis nigra, histamine-immuno-reactivity was restricted to cells and fibres of the nervous system. The occurrence of histamine or a related substance in the nervous system of flatworms, which represent primary bilateria, indicates the importance of this neuroactive substance in the animal kingdom.
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  • 48
    ISSN: 1432-0878
    Keywords: Fundic mucosa ; Stomach ; Pepsinogen ; Cell renewal ; Development, ontogenetic ; Immunocytochemistry ; Mouse (ICR)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Development and maturation of pepsinogen 1-producing cells were studied in the gastric fundic mucosa of the mouse by means of light- and electron-microscopic immunocytochemistry using rabbit anti-rat pepsinogen 1-serum. In the adult mouse, secretory granules in mucous neck cells, transitional mucous neck/chief cells and chief cells are immunolabeled. The numerical density of gold particles on zymogen granules is not significantly altered among different stages of maturation of chief cells. In addition, rough endoplasmic reticulum and Golgi complex of these cell types show a weak labeling. In mice from day 16 of gestation to postnatal day 14 mucous neck cells and chief cells cannot be distinguished, but only one type of pepsinogen 1-producing cell, called ‘primitive chief cell’, is identified in the fundic gland. The intensity of immunoreactivity of secretory granules in primitive chief cells is uniform within an individual cell but varies greatly among different cells. The majority of primitive chief cells contains weakly labeled granules regardless of the maturation stage of cells or of animals. On postnatal day 21, mucous neck, transitional and chief cells are distinguishable, and secretory granules in these cells are intensely immunolabeled as in the adult. These results suggest that pepsinogen 1-production rapidly increases with differentiation of mucouse neck and chief cells.
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  • 49
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    Cell & tissue research 260 (1990), S. 203-205 
    ISSN: 1432-0878
    Keywords: Immunocytochemistry ; Prolactin cells ; Pituitary gland ; Tilapia larvae, Oreochromis mossambicus (Teleostei)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Using an antiserum to highly purified chum salmon prolactin, prolactin cells were identified in the putative rostral pars distalis of newly hatched tilapia larvae (Oreochromis mossambicus) by the immunogold method for the electron microscope. In the putative rostral pars distalis, some cells had another kind of secretory granule which was much less numerous, much smaller in size, and without immunoreactivity to salmon prolactin antiserum. Controls incubated with salmon prolactin-preabsorbed antiserum or normal serum showed no immunoreactive cells, confirming the specificity of the antiserum. The possible role of prolactin in the osmoregulation of tilapia larvae is discussed.
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  • 50
    ISSN: 1432-0878
    Keywords: Antennae ; Motoneurons ; Immunocytochemistry ; Cobalt labelling ; GABA ; Gryllus bimaculatus (Insecta)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary In crickets, a deutocerebral motoneuron sends axon collaterals to 6 of the 7 antennal muscles. Previous results indicated that this neuron exerts inhibition on these muscles and thus may be a common inhibitory motoneuron. In our present study, we show by doublelabelling, i.e. retrograde cobalt-filling and GABA-immunocytochemistry, that this neuron is GABA-immunoreactive, thus demonstrating that one common inhibitory motoneuron is part of the antennal motor system of crickets.
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  • 51
    ISSN: 1432-0878
    Keywords: Neuropeptides ; Immunocytochemistry ; Submucosal plexuses ; Enteric nervous system ; Small intestine ; Pig
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary In addition to differences between the two submucosal ganglionic neural networks, i.e., the plexus submucosus externus (Schabadasch) and the plexus submucosus internus (Meissner), with respect to the occurrence and distribution of serotonin as neurotransmitter, immunocytochemistry also revealed a distinct distribution for various neuropeptides in these two plexuses. Immunoreactivity for galanin, vasoactive intestinal polypeptide, calcitonin gene-related peptide, substance P, neuromedin U, enkephalin, somatostatin and neuropeptide Y was found in varicose and non-varicose nerve fibres of both submucosal ganglionic plexuses, albeit with a distinct distributional pattern. The difference in neurotransmitter and/or neuromodulator content between both neural networks became even more obvious when attention was focussed on the immunoreactivity of the nerve cell bodies for these substances. Indeed, neuropeptide Y, enkephalin-and somatostatin-immunoreactive neuronal perikarya as well as serotonergic neuronal cell bodies appear solely in the plexus submucosus externus. Neuromedin U-immunoreactive perikarya, mostly coexisting with substance P, are observed in large numbers in the plexus submucosus internus, whilst they are rare in the plexus submucosus externus. Double-labelling immunostaining for substance P with CGRP and galanin revealed a different coexistence pattern for the two submucosal ganglionic plexuses. The differing chemical content of the neuronal populations supports the hypothesis that the existence of the two submucosal ganglionic plexuses, present in most large mammals including man, not only reflects a morphological difference but also points to differentiated functions.
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  • 52
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    Cell & tissue research 262 (1990), S. 177-190 
    ISSN: 1432-0878
    Keywords: FMRFamide ; Neuropeptide ; Immunocytochemistry ; Nervous system, central ; Neurohormones ; Helix pomatia (Mollusca)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The distribution of FMRFamide-like immunoreactive (FLI) neurons and their morphological characteristics have been investigated in the central nervous system of the snail, Helix pomatia L. Approximately phageal ganglion complex. More than 50% of the FLI neurons were located in the cerebral ganglia. The FLI neurons could be divided into four groups according to size: (i) giant neurons (over 100 μm); (ii) large neurons (80–100 μm); (iii) medium-sized neurons (40–70 μm); (iv) small neurons (12–30 μm). They were distributed i) in groups or clusters, typical of small neurons and ii) in solitary form or in groups comprising 2–3 cells, typical of large and giant neurons. Giant and large neurons revealed only limited arborizations in the neuropil, but rich branching towards and in the peripheral nerves. Some of the small neurons had extensive arborizations of varicose fibers in the neuropil. They may therefore play some role in integratory processes. Varicose FLI fibers were visualized in the cell body layer of the different ganglia, and in the neural sheath of both the ganglia and the peripheral nerves. We propose a multifunctional involvement of FLI neurons and FMRFamide-like neuropeptides in the Helix nervous system: (i) a synaptic or modulatory role in axo-axonic interactions in the neuropil; (ii) a direct influence on neuronal cell bodies in the cortical layer, (iii) innervation of different peripheral organs; and (iv) remote neurohormonal control of peripheral events through the neural sheath.
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  • 53
    ISSN: 1432-0878
    Keywords: Immunocytochemistry ; Gonadotropes ; Morphometry ; Stereology ; Rana pipiens (Anura)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Previous physiological results have indicated the existence of two releasable pools of gonadotropins in amphibian pituitaries: an acute releasable pool that appears independent of protein synthesis, and a storage pool involved in chronic release that depends on protein synthesis. To elucidate the ultrastructural localization of these pools and the morphological changes induced in gonadotrope cells after treatment with gonadotropin-releasing hormone, we carried out a morphometric study of immuno-identified gonadotrope cells using an in vitro superfusion system. Treatment with gonadotropin-releasing hormone induced a degranulation of small (110–255 nm) and medium (236–360 nm) secretory granules as well as hypertrophy of the endoplasmic reticulum and Golgi complex. Simultaneous incubation with gonadotropin-releasing hormone and cycloheximide inhibited the release of secretory granules although the endoplasmic reticulum and Golgi complex were hypertrophied. These morphological results strongly suggest: (1) that gonadotropin-releasing hormone induces degranulation and hypertrophy of the biosynthetic machinery in gonadotrope cells; and (2) that the activation of the endoplasmic reticulum and Golgi complex by stimulation with gonadotropin-releasing hormone is independent of protein synthesis, while the release of secretory granules is protein synthesis-dependent. In addition, the second or “storage” pool of gonadotropin is associated mainly with the small and medium secretory granules.
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  • 54
    ISSN: 1432-0878
    Keywords: Chang hepatoma cells ; Growth hormone ; GERL ; Golgi complex ; Immunocytochemistry ; Tumor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The binding and internalization of endogenous growth hormone in Chang hepatoma cells were localized on the cell surface and in the Golgi-endoplasmic reticulum-lysosome (GERL) area by various indirect immunocytochemical labeling techniques, namely, peroxidase or colloidal gold conjugated to secondary antibody, and avidin-biotin complex methods. Rabbit antiserum and monoclonal antibodies raised against HPLC-purified porcine growth hormone were used in this study. In fixed material, antigen-antibody complexes were found to be homogeneously distributed along the cell membrane. Control groups showed negative binding on the cell surface. Trypsin treatment before immunolabeling removed antibody binding completely, but hyaluronidase was ineffective. Pretreatment of lectins did not block the recognition of primary antibody to antigen molecules on cell surface. Internalization of the antigen-antibody peroxidase or gold complexes was demonstrated in the cells, which were immunolabeled at 4°C, and then reincubated for 0–30 min at 37°C before fixation. After reincubation, the internalized ligand complexes were found in vesicles near the cell surface or in the GERL area near the Golgi apparatus which, however, did not label for peroxidase. These findings suggest that the trypsin-sensitive growth hormone, specifically bound and internalized into Chang hepatoma cells, is localized in the GERL instead of the Golgi apparatus and might be involved in the mechanism of tumor cell growth.
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  • 55
    ISSN: 1432-0878
    Keywords: Galanin ; Immunocytochemistry ; Necturus maculosus (Urodela)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Galanin is a biologically active peptide which has a wide pattern of distribution in the mammalian central and peripheral nervous systems. However, the distribution of galanin-like immunoreactivity in amphibian species has not been well elucidated. In the present study, biochemical and immunohistochemical techniques were used to determine the relative concentrations, biochemical nature, and cellular localization of galanin-like immunoreactivity in the brain, heart, urinary bladder, and small intestine of Necturus maculosus (common name: mudpuppy). The results of this study indicate that each of these types of tissue contain a galanin-like peptide which is similar to porcine galanin. Brain and heart concentrations of galanin-like immuno-reactivity were particularly high, although substantial amounts were also present in the small intestine and urinary bladder. Galanin immunoreactivity was observed in ascending fiber tracts of the brainstem and in fibers in the hypothalamus. In addition, galanin immunoreactivity was observed in autonomic neurons and processes in the heart, bladder, and small intestine. The pattern of distribution of galanin-like immunoreactivity in many tissues of this amphibian species is similar to the previously described mammalian pattern; however, galanin-immunoreactive innervation of cardiac tissue has not been reported in mammals. We suggest that galanin-like immunoreactivity in the heart may be more extensive in amphibian species than in mammals.
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  • 56
    ISSN: 1432-0878
    Keywords: Immunocytochemistry ; Photoreceptor cells ; Rhodopsin ; Membrane recycling ; Cherax destructor (Crustacea)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Examination of the ultrastructure of retinula cells of the Australian crayfish Cherax destructor at different times over a 24-hour cycle, together with patterns of anti-rhodopsin antigenicity, has lead to the formulation of a model of photoreceptor membrane turnover in these animals. Its main features are: (a) the existence of two bursts of rhabdomeral membrane breakdown; one, light-sensitive and synchronous, occurring at dawn, the other, constituting the first part of the membrane replacement phase itself, occurring during the afternoon and night, (b) the desynchronisation of the replacement phase of turnover between animals and to a lesser extent between cells of the same retina, (c) confinement of ultrastructurally detectable signs of photoreceptor membrane processing to the retinula cells themselves, and (d) replacement of a substantial part if not all of the rhabdomeral membrane daily. This model is compatible with many of the observations reported on the American crayfish Procambarus, and utilises the same basic mechanisms that are believed to operate in photoreceptor membrane turnover in many other arthropod compound eyes.
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  • 57
    ISSN: 1432-0878
    Keywords: Pinealocytes ; Visinin ; Calcium-binding protein ; Light, constant ; Photosensitization ; Immunocytochemistry ; Domestic fowl (Gallus domesticus)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Visinin, a calcium-binding protein isolated from the soluble fraction of homogenized chick retinae, has been recognized immunocytochemically in the pinealocytes of various submammals. In the chick pineal organ, continuous environmental light induced an increase in population density of visinin-immunoreactive pinealocytes. In semi-quantitative, dot-immunoblotting analysis, the amount of visinin in the pineal organs of chicks kept under continuous light for 3 days was 4–8 fold more abundant than that under continuous darkness for the same duration. Eye-enucleation and organ culture experiments clarified that this lighting effect was exerted directly on the pineal organ through the skull, and not via the neural pathway including the retinohypothalamic projection. These data suggest the existence of direct photosensitivity in the chick pinealocyte itself and the possible involvement of visinin in photoreception of the pineal organ as well as the retina of chicks.
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  • 58
    ISSN: 1432-0878
    Keywords: Immunocytochemistry ; Vasotocin ; Hypothalamus ; Neurosecretory fibers ; Scyliorhinus canicula (Elasmobranchii)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The distribution of vasotocin-like peptides in the central nervous system of the cartilaginous fish Scyliorhinus canicula was determined by indirect immunofluorescence and peroxidase anti-peroxidase techniques, using a specific antiserum raised in rabbits against synthetic vasotocin. Immunoreactive perikarya were mainly detected in the anterior hypothalamus, within the midcaudal part of the preoptic nucleus. The most rostral positive cell bodies were located in the dorso-lateral parts of the preoptic area, whereas at a more caudal level, they took a ventro-medial position within the deepest layers of the nucleus. Throughout the preoptic region these cells varied in shape according to their location. Occasionally, scattered vasotocin-like immunopositive cells were also identified in the nucleus periventricularis hypothalami. Vasotocin immunoreactivity was detected in numerous varicose nerve fibers of the preopticohypophysial tract. These fibers were seen to course through the medio-basal hypothalamus and caudally, after having passed the hypophysial stem, they reached the neurointermediate lobe of the pituitary. Numerous immunoreactive fibers were also observed within the rostro-medial region of the median eminence. At this level the fibers were in close proximity to the capillary loops. In the preoptic region, some stained cells exibited short processes that appeared to contact non-reactive perikarya. By comparing the distribution of vasotocin- and corticotropin-releasing factor immunoreactivity on adjacent then serial sections, it was revealed that these peptides, in S. canicula, do not coexist in the same perikarya. The present results, are compared with those obtained in other vertebrate groups, and their possible functional implications are discussed.
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  • 59
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    Cell & tissue research 262 (1990), S. 543-550 
    ISSN: 1432-0878
    Keywords: PYY ; NPY ; CGRP ; Serotonin ; Lung ; Radioimmunoassay ; Immunocytochemistry ; Mesocricetus auratus (Rodentia)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary We investigated the presence of peptide YY, neuropeptide Y, calcitonin gene-related peptide and serotonin in the hamster lung by radioimmunoassay, high performance liquid chromatography and immunocytochemistry. Lung-tissue concentrations of peptide YY and neuropeptide Y were 1.3±0.2 and 2.5±0.2 pmol/g wet weight, respectively. These two closely related pancreatic peptides were demonstrated in separate peaks with high performance liquid chromatography. The peptide YY appeared fragmented as immunoreactive peptide YY eluted primarily late in the gradient but showed additional peaks early in the gradient. Peptide YY-like immunoreactivity (PYY-LI) was predominantly observed in one or more cells of neuroepithelial bodies in all airways peripheral to bronchioles, and in solitary neuroendocrine cells primarily located in the same peripheral areas. Neuropeptide Y-LI was seen in individual, thin nerve fibers around arteries and veins, in the airway lamina propria, and in the airway epithelium; in the latter also immunopositive nerve terminals were located. This pattern did not appear to coincide with that of calcitonin gene-related peptide-LI in epithelial nerve fibers and terminals. Peptide YY-LI, calcitonin gene-related-LI and serotonin-LI were present in cells of one and the same neuroepithelial body. However, peptide YY-LI was never found to be co-localized with calcitonin gene-related-LI or serotonin-LI, but the latter two were co-localized as previously reported.
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  • 60
    ISSN: 1432-0878
    Keywords: Hypothalamus ; Neurosecretion ; Vasotocin-neurophysin precursor ; Immunocytochemistry ; Lectin histochemistry ; Snake, Natrix maura ; Lizard, Liolaemus cyanogaster
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The immunocytochemical and lectin-binding properties of the magnocellular neurosecretory neurons in the hypothalamus of 2 reptilian species, the snake Natrix maura and the lizard Liolaemus cyanogaster, were investigated. Particular attention was paid to the secretory droplets present in these neurons. Antisera against bovine neurophysins I+II, arginine-vasotocin, and mesotocin were used. The following lectins were applied: concanavalin A (Con A), wheat-germ agglutinin (WGA), and Limax flavus agglutinin (LFA). Adjacent 1-μm-thick methacrylate sections were used to investigate the same secretory neuron and the same colloid droplets with all three antisera and all three lectins. Several sections were treated with trypsin and urea before immunostaining or lectin binding. Con A bound to both vasotocin- and mesotocin-immunoreactive neurons, WGA exclusively to vasotocin neurons; neither of these neurons reacted with LFA. The colloid droplets were present in vasotocin neurons but absent in the mesotocin neurons. These secretory droplets showed an affinity for Con A but not for WGA, and reacted with antisera against neurophysins and vasotocin. In Natrix maura, the colloid droplets became reactive with Con A and the antisera used only after pretreatment of the sections with trypsin and urea. Within the hypothalamo-neurohypophyseal system, antiserum against vasotocin and WGA revealed the same fiber bundles. It is concluded (i) that in reptiles the vasotocin-neurophysin precursor is glycosylated, (ii) that vasotocin neurons have the exclusive capacity to form colloid droplets, and (iii) that these droplets are an intracisternal (RER) storage form of the vasotocin-neurophysin precursor.
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  • 61
    ISSN: 1432-0878
    Keywords: Thyroid ; Neuromedin U ; C-cell ; Immunocytochemistry ; Chromatography ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Neuromedin U is a novel neuropeptide found to have a widespread distribution extending throughout the mammalian central nervous system, gastrointestinal tract and the endocrine cells of the pituitary gland. In order to investigate the possibility that neuromedin U-like immunoreactivity is also present in the thyroid gland of the adult rat we have examined its localisation and molecular nature by radioimmunoassay, immunocytochemistry and chromatographic analysis. The neuromedin U content of the whole thyroid gland was found to be 331±67 fmol/gland (mean±SEM), and this value significantly decreased (163±17 fmol/gland) as a result of 14 days of treatment with the anti-thyroid agent methimazole (10 mg/rat/day. Thyrotoxicosis induced by exogenous T4 (10 μg/rat/day) failed to alter the thyroid content of this peptide. Immunostaining studies localised neuromedin U to a minor population of parafollicular C-cells in untreated animals. Complementary chromatographic studies revealed a single molecular form of neuromedin U-like immunoreactivity in thyroid tissue extracts which was indistinguishable from synthetic rat neuromedin U standard.
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  • 62
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    Cell & tissue research 260 (1990), S. 41-48 
    ISSN: 1432-0878
    Keywords: Keratin filament ; Circumvallate papilla ; Taste bud ; Immunocytochemistry ; Electron microscopy ; Mouse (dd)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Keratin filaments of epithelial- and taste-bud cells in the circumvallate papillae of adult and developing mice were studied by immunocytochemistry using monoclonal antikeratin antibodies (PKK2 and PKK3) and by conventional electron microscopy. Elongated cells (type-I,-II, and-III cells) of the taste buds were stained by PKK3 antibody, which reacts with 45-kdalton keratin, whereas basal cells of the taste buds and surrounding epithelial cells showed negative staining with PKK3. Such PKK3-reactive cells occurred at 0 day after birth, when taste-buds first appeared in the dorsal surface epithelium of the papillae. Thus 45-kdalton keratin seems to be an excellent immunocytochemical marker for identifying taste-bud cells. Epithelial cells in all layers of the trench wall and basal layer cells of the dorsal surface contained densely aggregated bundles of keratin filaments that reacted with PKK2 antibody, but not with PKK3. In contrast, taste-bud cells and spinous and granular layer cells of the dorsal surface possessed loose aggregated bundles of filaments that reacted with PKK3, but not with PKK2. These results suggest that the aggregation and distribution pattern of keratin filaments may reflect differences in the keratin subtypes that comprise these filaments.
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  • 63
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    Cell & tissue research 261 (1990), S. 73-84 
    ISSN: 1432-0878
    Keywords: Muscle, skeletal ; Regeneration ; Thrombospondin ; Fibrinogen ; Injury ; Immunocytochemistry ; Rat (Sprague-Dawley)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Biochemical and immuno-microscopic techniques were used to study temporal involvement of thrombospondin in relation to fibrinogen in muscle regeneration using a rat skeletal muscle-wound model. In undamaged control muscle, no fibrinogen and minimal thrombospondin antigen was found. Following crushing injury, fibrin networks appear immediately, followed by a gradual ordered accumulation of thrombospondin (within a few hours) in the vicinity of the vascular bed and adjacent endomysial connective tissue. Later, thrombospondin becomes associated with connective tissue and basal laminae around muscle fibers throughout the damaged muscle, maximal labelling occurring 3–6 days post-injury. Thrombospondin immunoreactivity decreased thereafter to near normal levels after 7 days post-injury, coincident with the appearance of regenerating muscle fibers. In contrast, little fibrin material remained by five days after injury. Quantitative radioimmunoassay of soluble thrombospondin antigen and radioimmune labelling of thick frozen sections reinforced the qualitative immuno-microscopic observations, with levels peaking at 3–4 days post-trauma, 10-fold over control levels. SDS-PAGE immunoblotting of non-reduced muscle extracts three days after a crush assault shows that the bulk of the thrombospondin incorporated into the injury site exists in a polymerized state (≤1000 kD). These results demonstrate that the temporal appearance and disappearance of thrombospondin in the healing of a crushing lesion in muscle is related more closely to the regeneration phase of muscle than to the coagulation phase.
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  • 64
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    Cell & tissue research 261 (1990), S. 107-113 
    ISSN: 1432-0878
    Keywords: Dynorphin A ; GABA ; Substantia nigra ; Immunocytochemistry ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The opioid peptide dynorphin A (1–17) is the third transmitter identified in the striatonigral projection, the other two being gamma-aminobutyric acid (GABA) and substance P. The ultrastructural features of the dynorphinergic terminals in substantia nigra/pars reticulata were studied using pre-embedding immunocytochemistry with the classical peroxidase-antiperoxidase-diaminobenzidine-method; these features were compared with GABAergic boutons visualized with an immunogold method. Two distinct types of dynorphin-A-immunoreactive boutons could be identified: (1) type A (81%) possessing characteristics similar to the GA-BAergic nerve endings in this region, i.e., large pleomorphic vesicles and symmetric synaptic contacts, (2) type B (19%) displaying asymmetric synaptic zones and small, mostly round vesicles. These results are in agreement with physiological studies suggesting a dual action of dynorphin A in substantia nigra.
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  • 65
    ISSN: 1432-0878
    Keywords: Neuropeptides ; FMRFamide ; Immunocytochemistry ; Insect nervous system ; Manduca sexta (Insecta)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Using an antiserum against the tetrapeptide FMRFamide, we have studied the distribution of FMRFamide-like substances in the brain and suboesophageal ganglion of the sphinx mothManduca sexta. More than 2000 neurons per hemisphere exhibit FMRFamide-like immunoreactivity. Most of these cells reside within the optic lobe. Particular types of FMRFamide-immunoreactive neurons can be identified. Among these are neurosecretory cells, putatively centrifugal neurons of the optic lobe, local interneurons of the antennal lobe, mushroom-body Kenyon cells, and small-field neurons of the central complex. In the suboesophageal ganglion, groups of ventral midline neurons exhibit FMRFamide-like immunoreactivity. Some of these cells have axons in the maxillary nerves and apparently give rise to FMRFamide-immunoreactive terminals in the sheath of the suboesophageal ganglion and the maxillary nerves. In local interneurons of the antennal lobe and a particular group of protocerebral neurons, FMRFamide-like immunoreactivity is colocalized with GABA-like immunoreactivity. This suggests that FMRFamide-like peptides may be cotransmitters of these putatively GABAergic interneurons. All FMRFamide-immunoreactive neurons are, furthermore, immunoreactive with an antiserum against bovine pancreatic polypeptide, and the vast majority is also immunoreactive with an antibody against the molluscan small cardioactive peptide SCPB. Therefore, it is possible that more than one peptide is localized within many FMRFamide-immunoreactive neurons. The results suggest that FMRFamide-related peptides are widespread within the nervous system ofM. sexta and might function as neurohormones and neurotransmitters in a variety of neuronal cell types.
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  • 66
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    Cell & tissue research 261 (1990), S. 367-373 
    ISSN: 1432-0878
    Keywords: Angiotensinogen ; Ovary ; Estrous cycle Renin-angiotensin system ; Atresia ; Immunocytochemistry ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The present study examined the presence and cellular distribution of angiotensinogen, the precursor to the angiotensin peptides, in the ovary of the normal cycling rat by immunocytochemistry. Angiotensinogen staining was present in the granulosa cells of maturing follicles and to a lesser extent in those undergoing atresia. Staining was not seen in the granulosa cells of primordial or early primary follicles. In maturing follicles intense staining for angiotensinogen was confined to the antral cell layers, cells of the cumulus oophorus and in the follicular fluid. Strong immunostaining was also seen in the germinal epithelium covering the ovary. Lighter angiotensinogen staining was observed in some parts of the cortical and medullary stroma and occasionally in corpora lutea. No variation in the intensity or pattern of angiotensinogen staining was observed throughout the estrous cycle. Comparison of the distribution of angiotensinogen with the previously described localization of renin, AII, angiotensin converting enzyme and AII receptors, suggests that there are a number of intra-ovarian sites at which AII could be produced.
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  • 67
    ISSN: 1432-0878
    Keywords: Chorionic gonadotropin ; Placental lactogen Placenta ; Cryoultrastructure ; Immunocytochemistry ; Human
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The sites of intracellular synthesis and storage of human placental lactogen (hPL) and human chorionic gonadotropin (hCG) are controversial. We have used one of the most sensitive methods, cryoultramicrotomy and immunogold labelling, to localise these hormones at the electron-microscopic level. In both 12-week and term placentas hCG and hPL are present throughout the rough endoplasmic reticulum cisternae, in the Golgi bodies, and in the infrequent small dense granules of the syncytiotrophoblast. Previous assays have shown that hCG is at a higher concentration in early pregnancy and hPL peaks in late pregnancy, and our results corroborate these findings. No significant localisation of either hormone was seen in the cytotrophoblast or villous stroma. The results suggest that both hCG and hPL are synthesised and packaged by the classical secretory pathway, although the level of hormone stored in granules at any one time is small.
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  • 68
    ISSN: 1432-0878
    Keywords: GRF ; Immunocytochemistry ; Brain ; Pituitary ; Hypothalamus ; Vasotocin ; Anguilla anguilla ; Carassus auratus (Teleostei)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary An antiserum to growth hormone-releasing factor (GRF) 1-44 was applied on brain and pituitary sections of nine teleost species. Immunoreactive (ir) perikarya were demonstrated in parvo- and magnocellular portions of the preoptic nucleus (PON) and occasionally in the nucleus lateralis tuberis. The two tracts originating in the PON ran ventro-laterally toward the optic chiasm and then caudally in the basal hypothalamus. In the pars distalis (PD) of the eel, carp, goldfish and salmonids, GRF-ir fibers did not enter the rostral PD and few fibers passed close to somatotropes. In.Myoxocephalus andMugil, a variable number of ir-fibers passed close to cells of the rostral and proximal PD. In the neurointermediate lobe, GRF-ir fibers were located exclusively in the neural tissue of the eel and trout. In goldfish, carp andMyoxocephalus, GRF-ir fibers entered the intermediate lobe. This antiserum also labeled corticotrops and, to a lesser extent, melanotrops in the pituitary of cyprinids. A variable number of perikarya contained both GRF and vasotocin in the PON of the eel. In all teleost species studied so far, the distribution patterns of GRF are different, and the function of the various adenohypophysial cell types appears to be differently modulated, according to the variable distribution of GRF in the pituitary.
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  • 69
    ISSN: 1432-0878
    Keywords: Bombesin-like peptide ; Immunocytochemistry ; Hypothalamus ; Neurosecretory ; Fibers ; Scyliorhinus canicula (Elasmobranchii)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The presence and distribution of bombesin-like material were investigated in the brain of the cartilaginous fishScyliorhinus canicula using conventional immunocytochemical techniques. Perikarya containing bombesin-like immunoreactivity were identified in the hypothalamus, within the magnocellular component of the preoptic nucleus. Some immunopositive elements appeared to be of cerebrospinal fluid-contacting type. Beaded immunoreactive fibers were seen crossing the ventral telencephalon and the whole hypothalamus. An important tract of fibers was found in the infundibular floor and in the median eminence, in close contact with the vascular system of the pituitary portal plexus. A moderate number of positive fibers innervated the habenular complex and the dorsal wall of the posterior tuberculum. These findings indicate that a neuropeptide strictly related to amphibian bombesin is located in specific hypothalamic neurons ofS. canicula. The distribution of the immunoreactive fibers and terminals suggests that, in fish, this peptide, may be involved in neuroendocrine and neuromodulator functions.
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  • 70
    ISSN: 1432-0878
    Keywords: Immunocytochemistry ; Development, ontogenetic ; Serotonin-like immunoreactivity ; identified neurons ; cerebral ganglion ; Tenebrio molitor (Insecta)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Serotonin-immunoreactive neurons in the brain of Tenebrio molitor L. have been demonstrated and mapped throughout metamorphosis. Most serotonin-immunoreactive brain neurons persist throughout metamorphosis; their fate can be followed during development because of their characteristic cell body locations and arborization patterns. The detailed morphology of the persisting neurons, however, changes during metamorphosis, probably to accommodate architectural changes of the different brain centers. Serotonin-immunoreactivity in the optic lobes allows a subset of neurons that is newly differentiated during metamorphosis to be identified. Phylogenetic homology of serotonin-immunoreactive brain interneurons of different insect species is discussed. The serotonin-immunoreactive brain neurons comprise a phylogenetically conserved neuronal population. Serial homologous abdomino-thoracic and brain serotonin-immunoreactive neurons were characterized, allowing a comparison of some basic structural features of these neurons.
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  • 71
    ISSN: 1432-0878
    Keywords: Cerebrospinal fluid-contacting neurons ; Paraventricular organ ; Posterior recess organ ; Somatostatin ; Serotonin ; Ultrastructure ; Immunocytochemistry ; Dogfish,Squalus acanthias skate,Raja radiata (Elasmobranchii)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The paraventricular organ (PVO) and the posterior recess organ (PRO) of two elasmobranch species, the spiny dogfish,Squalus acanthias, and the skate,Raja radiata, were investigated by use of scanning and transmission electron microscopy and immunocytochemistry employing a series of primary antisera. The PVO and PRO contained four types of cerebrospinal fluid (CSF)-contacting neurons. One type was free of secretory granules and projected a dendrite-like process into the ventricle. The other three types were distinguished according to the size of their secretory granules. The ventricular extensions of these cells were filled with secretory granules. By means of immunocytochemistry three types of CSF-contacting neurons were observed in the PVO and PRO. Type I contained only serotonin; type 2 displayed only somatostatin; type 3 was endowed with both serotonin and somatostatin. Type I dominated in the PRO, whereas type 3 was the most frequent in the PVO. The latter cells appear to be the site of origin of a loose tract formed by serotonin- and somatostatinimmunoreactive fibers projecting from the PVO into the neuropil of the PRO. Compact bundles formed exclusively by serotonin fibers were also shown to extend between the PVO and PRO. The basal processes of the CSF-contacting neurons of the PRO penetrated into the underlying neuropil. This neuropil is rich in synapses and can be regarded as an integrative area to which the basal processes of the local CSF-contacting neurons, serotonin and somatostatin fibers from the PVO, and fibers containing immunoreactive thyrotropin-releasing hormone of unknown origin, support a conspicuous input. The present findings indicate that the PVO and PRO of elasmobranchs are functionally integrated structures.
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  • 72
    ISSN: 1432-0878
    Keywords: Oxytocin ; Corpus luteum ; Luteinized cyst ; Immunocytochemistry ; Ewe (Romney Marsh)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Anoestrous Romney Marsh ewes with or without progesterone pretreatment were injected with multiple low-doses of gonadotrophin-releasing hormone followed by a single, larger bolus. Blood samples were taken at twelve-hourly intervals for progesterone radioimmunoassay. Ewes were slaughtered on day 3 or 5 after the bolus injection, and the ovaries were collected for histology and immunocytochemical examination for oxytocin-immunocreactivity. The corpora lutea of all ewes killed on day 3 had similar weights and morphology. The ovaries of those ewes which were not pretreated with progesterone also contained some luteinized cysts. Ewes slaughtered on day 5 were separated into 2 groups according to plasma progesterone profiles, which were either rising (‘normal’), or falling after a transitory rise (‘abnormal’). Those ewes pretreated with progesterone all had a ‘normal’ progesterone profile whereas, of 14 ewes not pretreated with progesterone, 6 were ‘normal’ and 8 ‘abnormal’. Corpora lutea were significantly lighter in the ‘abnormal’ group and the ovaries of most of these ewes also contained luteinized cysts. All corpora lutea and luteinised cysts showed staining for oxytocin-immunoreactivity although the staining intensity was variable. In corpora lutea from ‘normal’ ewes oxytocin was restricted to large luteal cells. In addition tissues from ‘abnormal’ ewes also contained many cells with an atypical elongated shape which stained for oxytocin-immunoreactivity. These results show that progesterone pretreatment is needed for both normal morphological and endocrine development of corpora lutea in anoestrous ewes stimulated with gonadotrophin-releasing hormone.
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  • 73
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    Cell & tissue research 262 (1990), S. 569-577 
    ISSN: 1432-0878
    Keywords: Blastocyst ; Uterine proteins ; Uteroglobin ; Endocytosis ; Autoradiography ; Immunocytochemistry ; Acid phosphatase ; Rabbit
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Uptake of uteroglobin (UGL) by day-6 rabbit blastocysts and the intracellular fate of this protein were studied by light- and electron-microscopic autoradiography, immunocytochemistry and acid-phosphatase cytochemistry. UGL, labelled with N-succinimidyl-(2-3-3H)-propionate, was administered to embryos in vitro for 15 min to 4 h. The kinetics, determined from light-microscopic autoradiographs, showed a continuous uptake of the labeled protein over a 4-h period of incubation. At the ultrastructural level, increasing numbers of silver grains and an intense UGL immunoreaction in protein vacuoles and crystalloid bodies of trophoblast cells indicated that 3H-UGL had accumulated in these organelles. The presence of crystalloid inclusions in protein vacuoles suggests their origin by a condensation of the protein content, including UGL. Lysosomes containing radioactivity were rarely found, suggesting a very low degradation rate of the 3H-UGL. Protein vacuoles and crystalloid bodies exhibited no acid phosphatase reaction. The enzyme was mainly found outside the basal and lateral cell membranes of trophoblast cells, and on the rough endoplasmic reticulum of endoderm cells.
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  • 74
    ISSN: 1432-0878
    Keywords: Median eminence ; Catecholamines ; Neuropeptides ; Immunocytochemistry ; Double labeling ; Ultrastructure ; Triturus alpestris (Urodela)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Dopaminergic and peptidergic nerve fibers were simultaneously demonstrated with a double-labeling technique at the ultrastructural level. The first antibody, raised against tyrosine hydroxylase, was applied during the preembedding phase and visualized with the peroxidase method. The second antibody, raised against one of the peptides met-enkephalin, somatostatin or gonadotropin-releasing hormone (GnRH), was applied to the ultrathin sections and visualized with gold-labeled goat anti-rabbit IgG. The fibers of both categories were present in the zona externa of the median eminence, frequently contacting the basal lamina of the portal vessels. In addition, topographical relationships between different types of nerve fibers were observed in the perivascular areas, although there were no morphological signs of synaptic specializations. Using serial sections, it could be established that one GnRH-fiber contacted both a dopaminergic fiber and a fiber immunoreactive for met-enkephalin. The observations support earlier physiological data concerning the regulation of the hypothalamo-hypophyseal axis, with special emphasis on the release of neurohormones in the median eminence of the newt.
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  • 75
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    Cell & tissue research 260 (1990), S. 381-386 
    ISSN: 1432-0878
    Keywords: Molluscan insulin-related peptide ; Neuropeptide ; Immunocytochemistry ; In situ hybridization ; Lymnaea stagnalis (Mollusca) ; Aplysia californica (Mollusca)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary the occurrence of insulin-related substances in the central nervous system of pulmonates and Aplysia californica was investigated by means of immunocytochemistry and in situ hybridization. Previous experiments have shown that, in Lymnaea stagnalis, the growth hormone-producing neurons in the cerebral ganglia (the so-called light green cells) express at least 5 genes that are related to the vertebrate insulin genes, i.e., they encode prohormones that are composed of a B- and A-chain and a connecting C peptide. These insulin related molecules also have the amino acids essential for their tertiary structure (viz. cysteines) at identical positions to those of the vertebrate insulins. In the investigated basommatophoran and stylommatophoran snails and slugs, neurons reacted with an antiserum raised against the C peptide of one of the molluscan insulin-related peptides. These neurons can be considered to be, based on morphological and endocrinological criteria, homologous to the light green cells of L. stagnalis. In A. californica, all central ganglia contain immunoreactive neurons. The highest number (about 50) was observed in the abdominal ganglion. The present results indicate that insulin-related substances are generally occurring neuropeptides in the central nervous system of molluscs.
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  • 76
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    Cell & tissue research 262 (1990), S. 105-113 
    ISSN: 1432-0878
    Keywords: Subcommissural organ ; Lectin histochemistry ; Immunocytochemistry ; Glycoproteins ; Human fetuses ; Bovine
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The subcommissural organ (SCO) of 7 human fetuses, 3 to 6.5 months old, was investigated by means of: (i) immunocytochemistry employing three different antisera against secretory products extracted from the bovine SCO and Reissner's fiber; (ii) lectin binding using concanavalin A (Con A; affinity: mannose, glucose), wheat-germ agglutinin (WGA; affinity: N-acetyl-glucosamine, sialic acid), and Limax flavus agglutinin (LFA; affinity: sialic acid). Sections of bovine SCO were processed simultaneously and examined for comparative purposes. The human fetal SCO displayed lectin-binding properties identical to those in the SCO of other mammals. Thus, Con-A-binding sites were restricted to abundant supranuclear structures that most likely corresponded to the rough endoplasmic reticulum, but were missing from granules located in the apical cytoplasm. The latter secretory material was strongly WGA- and LFA-positive and formed a distinct zone in the most apical portion of the ependymal cells. In contrast, this type of reactivity was missing in the adjacent cells of ependyma proper. In the bovine SCO, LFA-positive granules were also aggregated in an apical layer. The secretory material in the bovine SCO, especially its apical granular component, was strongly immunoreactive with the three antisera used; the human fetal SCO, however, lacked this immunoreactivity. It is postulated that the SCO of human fetuses secretes glycoproteins with a carbohydrate chain similar to-and a protein backbone different from-the secretions elaborated by the SCO of other vertebrate species.
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  • 77
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    Cell & tissue research 260 (1990), S. 431-434 
    ISSN: 1432-0878
    Keywords: Actin ; Cytoskeleton ; Immunocytochemistry ; Photoreceptor cells ; Drosophila melanogaster (Insecta)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The Drosophila ninaC mutation produces small rhabdomeres with the axial filament of the microvillar cytoskeleton reduced or missing. Using post-embedding immunogold labelling of LR White-embedded eyes, we show that several alleles of this mutation retain positive anti-actin immunoreactivity in the rhabdomeres, comparable to that of wild-type flies.
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  • 78
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    Cell & tissue research 261 (1990), S. 461-466 
    ISSN: 1432-0878
    Keywords: Secretion ; Plasmalemma ; Membrane dynamics ; Immunocytochemistry ; Freeze-fracturing ; Rat (Wistar)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The movements of the molecular components of the luminal plasma membrane during exocytotic secretion in parotid acinar cells were examined. For immunocytochemical study, we used an antiserum of dipeptidyl peptidase IV as a marker for the components of the luminal plasma membrane of acinar cells. In unstimulated acinar cells, dipeptidyl peptidase IV immunoreactivity is restricted to the luminal plasma membrane. However, after secretion was stimulated with a β-adrenergic agonist, isoproterenol, immunostaining became detectable on the membrane of discharged granules. Freeze-fracture images showed that the density of intramembrane particles on the P-fracture leaflets of discharged granule membranes is much higher than that of undischarged granule membranes during secretion. These results suggest that in parotid acinar cells of the rat, the components of the luminal plasma membrane move laterally, during secretion, to the membranes of discharged granules.
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  • 79
    ISSN: 1432-0878
    Keywords: Serotonin (5-HT) ; Substance P ; GABA ; Immunocytochemistry ; Invertebrate peripheral nervous system ; Mytilus galloprovincialis (Mollusca)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Immunocytochemical methods were applied to study the distribution of putative neurotransmitters (5-HT, substance P, GABA, glutamate and aspartate) in the nerve plexuses of the foot and the anterior byssus retractor muscle (ABRM) of Mytilus galloprovincialis (Mollusca, Bivalvia). The foot presents extensive nerve plexuses containing 5-HT and substance P-like immunoreactive material with a similar distribution beneath the surface epithelium, around the vessels and in the glandular regions. Coexistence of the two putative neurotransmitters was observed in a few nerve fibers, Conversely, muscle fibers, both in the foot and in the ABRM, are innervated only by 5-HT-positive fibers, while substance P-like material is present only in the networks of the ABRM epimysial sheath. Immunoreactivity for glutamate and aspartate was not demonstrated, while rare GABA-positive nerve cells and fibers were found only in the foot. The results of this investigation provide a morphological background to previous physiological studies on 5-HT in the nervous system of bivalve molluscs. Moreover, they confirm that the nervous system of Mytilus contains a remarkable amount of a substance related to the vertebrate tachykinin family.
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  • 80
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    Cell & tissue research 261 (1990), S. 517-524 
    ISSN: 1432-0878
    Keywords: Adrenal cortex ; Adrenal medulla ; Somatostatin immunoreactivity ; Immunocytochemistry ; Hybridization, in situ ; Rat
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Previous studies have shown that somatostatin modulates angiotensin-induced aldosterone secretion by adrenal glomerulosa cells. This effect is mediated through specific receptors which do not show any preference for somatostatin-14 (S14) or the N-extended form somatostatin-28 (S28). The study of the distribution of 125I-Tyr [Tyr0, DTrp8] S14-and 125I-Tyr[Leu8, DTrp22, Tyr25] S28-binding in frozen sections of the rat adrenal by autoradiography indicated that both peptides bind to similar loci. High concentrations of binding sites were observed in the zona glomerulosa, and low concentrations were detected in the medulla. At the ultrastructural level, immunocytochemistry after cryoultramicrotomy revealed endogenous S14-and S28-like immunoreactive material in zona glomerulosa and in medulla. In glomerulosa cells, immunoreactive material was localized at the plasma membrane level, in the cytoplasmic matrix, in the mitochondria, and in the nucleus. S14-and S28-like materials were detected in both epinephrine and norepinephrine-storing cells of the adrenal medulla. In these cells, the distribution of either immunoreactive product was similar; it was observed in cytoplasmic matrix, secretory granules and nucleus, but not at the plasma membrane level. In situ hybridization does not reveal somatostatin mRNA in zona glomerulosa or medulla. These results demonstrate that S14 and S28 bind to, and are taken up by zona glomerulosa and adrenal medullary cells, but are not produced by these cells.
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  • 81
    ISSN: 1432-0878
    Keywords: Neuropeptides (pancreatic polypeptide, peptide YY, neuropeptide Y) ; Immunocytochemistry ; Confocal scanning laser microscopy ; Schistosoma mansoni (Scolecida, Trematoda)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The presence and distribution of neuropeptides belonging to the pancreatic polypeptide family have been demonstrated by an indirect immunofluorescence technique in the nervous systems of adult male and female Schistosoma mansoni. Seven antisera of differing regional specificity to pancreatic polypeptide (PP), peptide YY (PYY) and neuropeptide Y (NPY) were employed on both whole-mount and cryostat-sectioned material. Positive immunoreactivity (IR) was obtained with all antisera except an N-terminally-directed antiserum to NPY. In the CNS, immunoreactivity was restricted to cell bodies and nerve fibres in the anterior ganglia, central commissure and dorsal and ventral nerve cords of both sexes, whereas, in the PNS, positive-IR was present in the plexuses innervating the subtegumental musculature and the oral and ventral suckers. Intense immunoreactivity was observed in a plexus of nerve fibres and cell bodies in the lining of the gynaecophoric canal and in fine nerve fibres innervating the dorsal tubercles of the male. In contrast, in the female, strong immunoreactivity was evident in nerve plexuses innervating the lining of the ovovitelline duct and in the wall of the ootype, but most notably in a cluster of cells in the region of Mehlis' gland. Results suggest that molecules with C-terminal homology to the PP-family are present in S. mansoni. These peptides would appear to be important regulatory molecules in the parasite's nervous system and may play a role in the control of egg production.
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  • 82
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    Cell & tissue research 262 (1990), S. 115-123 
    ISSN: 1432-0878
    Keywords: ACTH ; Immunocytochemistry ; Brain ; Hypothalamus ; Corticotropin-releasing factor ; Pituitary ; Carassius auratus, Salmo fario, Anguilla anguilla (Teleostei)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Immunostaining of brain and pituitary sections of three teleost species (goldfish, trout and eel) with antisera to porcine and human ACTH 1–39 revealed the presence of an ACTH (adrenocorticotropic hormone)-like peptide in the ventral hypothalamus. Perikarya were localized in the rostral, median and posterior portions of the nucleus lateralis tuberis (NLT); some were in contact with the cerebrospinal fluid. A dense network of immunoreactive (ir) fibers occurred in the peri-infundibular region and extended into the periventricular tissue, around the lateral and posterior recesses. Rostrally directed ir-fibers reached the telencephalon either ventrally or mediodorsally; some were observed in the olfactory lobe. In the mesencephalon, ir-fibers penetrated into the optic tectum of the goldfish. In the pituitary, both antisera intensely labeled rostral ACTH cells. Small groups of labeled cells were scattered in the rostral pars distalis and the proximal pars distalis. A gradient of activity was evident among ACTH cells: those located along the rostral neurohypophysis containing corticotropin-releasing factor nerve terminals were larger and often more marked than those farther away from the neural tissue. ACTH-like peptide in the brain may act as a neuromodulator, mainly in the NLT and the preoptic nucleus, and around the nuclei of the ventricular recesses containing serotonin and catecholamines.
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  • 83
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    Protoplasma 158 (1990), S. 109-120 
    ISSN: 1615-6102
    Keywords: Differentiation ; Immunocytochemistry ; [3H]T-autoradiography ; Stem cells ; Turbellaria
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A combination of microscopical, immunocytochemical, and autoradiographic techniques were employed to study stem cells and their fates during asexual reproduction and regeneration in two microturbellarians,Microstomum lineare (Macrostomida) andStenostomum leucops (Catenulida). Special attention was paid to the development of the immunoreactivity (IR) to FMRF/RF-amide and 5-HT in differentiating nerve cells. Asexual reproduction inM. lineare andS. leucops occurs by paratomy, i.e., fragmentation after completed differentiation of the new organs. Regeneration, on the other hand, involves a combination of morphallactic and epimorphic processes without the formation of a regeneration blastema. The only cells incorporating tritiated thymidine ([3H]T) were the mesenchymal and gastrodermal neoblasts, which proliferate continuously replenishing the population of stem cells available for growth, asexual reproduction and regeneration. These proliferative cells occurred in two ultrastructurally different forms, differing from each other only by the presence or absence of ciliar basal bodies in the cytoplasm. Few differentiated cells were labeled in the head piece after completed regeneration. A greater amount of labeled differentiated cells were, however, observed postpharyngeally in the first zooid as well as in zooids having developed during the same time (i.e., 20–45 h after the treatment with [3H]T). Furthermore, many labeled cells were still undifferentiated at that time or just in the beginning of the differentiation process. It can therefore be concluded that neoblasts function both as reserve cells and as functional stem cells for all differentiated cell types in these worms. IR to FMRF/RF-amide neuropeptides was not observed in nerve cells differentiating from neoblasts until the occurrence of dense-core vesicles in their cytoplasm. Due to methodological difficulties only weak or no IR to 5-HT could be traced in the nervous system of the asexual and regenerating worms.
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  • 84
    ISSN: 1615-6102
    Keywords: α-Amylase isozymes ; Barley aleurone ; Endoplasmic reticulum ; Golgi apparatus ; Immunocytochemistry ; Intracellular transport
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The localization of α-amylase (EC 3.2.1.1) in barley (Hordeum vulgare L. cv Himalaya) aleurone protoplasts was studied using electron microscope immunocytochemistry. Antibodies were raised against total barley α-amylase, i.e., α-amylase containing both highisoelectric point (high-pI) and low-pI isoforms, as well as against purified high- and low-pI isoforms. All antibodies localized α-amylase to the endoplasmic reticulum (ER) and Golgi apparatus (GApp) of the aleurone cell, and various controls showed that the labeling was specific for α-amylase. Labeling of protein bodies and spherosomes, which are the most abundant organelles in this cell, was very low. There was no evidence that α-amylase isoforms were differentially distributed within different compartments of the endomembrane system. Rather, both high- and low-pI isoforms showed the same pattern of distribution in ER and in the cis, medial, and transregions of the GApp. We conclude that in the Himalaya cultivar of barley, all isoforms of α-amylase are transported to the plasma membrane via the GApp.
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    Plant and soil 126 (1990), S. 115-119 
    ISSN: 1573-5036
    Keywords: inorganic P ; organic P ; P/Fe ; P/Mn ratios ; phosphorus ; rice ; silicon
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract A pot experiment was conducted to measure the effect of silicon on phosphorus uptake and on the growth of rice at different P levels. Rice (Oryza sativa L. cv. Akebono) was cultured in Kimura B nutrient solution without and with silicon (1.66 mM Si) and with three phosphorus levels (0.014 mM P, low; 0.21 mM, medium; and 0.70 mM, high). Shoot dry weight with Si (+Si) in solution increased with increasing P level, while shoot weight without Si (−Si) was maximum at 0.21 mM P, suggesting that +Si raised the optimum P level for rice. +Si increased shoot weight more when P was low or high than when P was medium. The concentration and amount of inorganic P in shoots increased with increasing P level. +Si did not significantly decrease P uptake by rice at 0.014 mM P, however, uptake at 0.21 and 0.70 mM P was 27 and 30 percent less than uptake with −Si, respectively. In −Si with 0.21 and 0.70 mM P, inorganic P in shoots was more than double the concentration in shoots grown in +Si solutions. The Si concentration in shoots decreased slightly with increasing P level, although Si uptake was not significantly affected by P. +Si decreased the uptake of Fe and Mn by an average of 20 and 50 percent, respectively, thus P/Mn and P/Fe ratios increased in the shoot when P was low. From the results above, the beneficial effect of Si on the growth of rice was clearly shown when P was low or high. This effect may have resulted from decreased Mn and Fe uptake, and thus increased P availability within P deficient plants, or from reduced P uptake when P was high.
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    Plant and soil 126 (1990), S. 121-125 
    ISSN: 1573-5036
    Keywords: P adsorption ; P desorption ; P/Mn ratio ; rice ; silicic acid ; Yakuno soil
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract A pot experiment was conducted to analyze the effect of silicon on the growth of rice grown in a P-deficient soil and on P availability in the soil. Silicic acid was used, rather than a silicate salt, to avoid the complication of changes in soil pH. Shoot dry weight on silicic acid treated soil (0.47 mg Si g−1 soil) increased significantly under both nonflooded and flooded conditions. Shoot Si concentration also increased although P concentration did not. Mn concentration decreased with silicic acid, resulting in a higher P/Mn ratio in shoots. An adsorption and desorption experiment showed that silicic acid did not displace P nor decrease the ability of the soil to adsorb P. In contrast, Si desorption increased with increasing P concentration in the solution, and Si adsorption was reduced when P was applied first. These results suggest that silicic acid does not increase P availability in soil. Increased dry weight may be attributed to a higher P/Mn ratio in the shoot, which may improve P utilization in the plant.
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    Plant and soil 124 (1990), S. 221-225 
    ISSN: 1573-5036
    Keywords: maize ; pollen fertility ; Zea mays L. ; zinc
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Zinc deficiency decreased pollen viability in maize (Zea mays L. cv. G2) grown in sand culture. On restoring normal zinc supply to zinc-deficient plants before the pollen mother cell stage of anther development, the vegetative yield of plants and pollen fertility could be recovered to a large extent, but the recovery treatment was not effective when given after the release of microspores from the tetrads. If zinc deficiency was induced prior to microsporogenesis it did not significantly affect vegetative yield and ovule fertility, but decreased the fertility of pollen grains, even of those which visibly appeared normal. If the deficiency was induced after the release of microspores from the tetrads, not only vegetative yield and ovule fertility but pollen fertility also remained unaffected.
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  • 88
    ISSN: 1573-5036
    Keywords: maize ; nutrient concentration ; nutrient availability ; root growth ; root zone temperature ; shoot growth ; shoot meristem temperature
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Maize seedlings were grown for 10 to 20 days in either nutrient solution or in soils with or without fertilizer supply. Air temperature was kept uniform for all treatments, while root zone temperature (RZT) was varied between 12 and 24°C. In some treatments the basal part of the shoot (with apical shoot meristem and zone of leaf elongation) was lifted up to separate the indirect effects of root zone temperature on shoot growth from the direct effects of temperature on the shoot meristem. Shoot and root growth were decreased by low RZT to a similar extent irrespective of the growth medium (i.e. nutrient solution, fertilized or unfertilized soil). In all culture media Ca concentration was similar or even higher in plants grown at 12 as compared to 24°. At lower RZT concentrations of N, P and K in the shoot dry matter decreased in unfertilized soil, whereas in nutrient solution and fertilized soil only the K concentration decreased. When direct temperature effects on the shoot meristem were reduced by lifting the basal part of the shoot above the temperature-controlled root zone, shoot growth at low RZT was significantly increased in nutrient solution and fertilized soil, but not in unfertilized soil. In fertilized soil and nutrient solution at low RZT the uptake of K increased to a similar extent as plant growth, and thus shoot K concentration was not reduced by increasing shoot growth rates. In contrast, uptake of N and P was not increased, resulting in significantly decreased shoot concentrations. It is concluded that shoot growth at suboptimal RZT was limited both by a direct temperature effect on shoot activity and by a reduced nutrient supply through the roots. Nutrient concentrations in the shoot tissue at low RZT were not only influenced by availability in the substrate and dilution by growth, but also by the internal demand for growth.
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  • 89
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    Plant and soil 126 (1990), S. 227-235 
    ISSN: 1573-5036
    Keywords: Oryza sativa ; rice ; root length density ; soil impedance ; tillage ; water stress
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The influence of various tillage methods on two wetland rice soils in the Philippines is reported. The soils differed principally in clay content, 38% for the clay loam (clayey, mixed isohyperthermic Entic Hapludoll) while 56% for the clay (clayey, mixed noncalcareous, isohyperthermic Andaqueptic Haplaquoll). This had a marked effect on their response to tillage and varying water regime. The clay soil, under field conditions, showed little change in pore size distribution or soil water behaviour with different tillage methods. Crop (Rice, Oryza sativa L., var. IR20) yields were unaffected by tillage. In contrast, tillage effects were very marked in the clay loam soil, which consisted of a greenhouse and a field trial. In the greenhouse, which experienced severe dry periods, wet tillage not only increased the moisture retentivity but also the soil impedance at soil matric potential (ψ)〈−0.01 MPa. Seasonal average ψ was 〈−1 MPa. Root length density decreased by 39% with dry tillage and by 56% with wet tillage compared with zero tillage. Grain yield however, did not vary with soil treatment. In the field, which experienced moderate dry spells, ψ varied between −0.13 and −0.48 MPa. Root length density was significantly reduced at soil impedance 〉0.75 MPa. Wet tillage increased soil moisture storage which minimized the soil impedance during the dry cycle more effectively than did dry tillage. The crop performed best under wet tillage and least under zero tillage. Wet tillage in this soil was more effective under moderate than under severe water stress conditions.
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  • 90
    ISSN: 1573-9104
    Keywords: soil salinity ; grain quality ; rice ; protein
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Four varieties of rice, differing in salinity tolerance and grown in saline soil (electrical conductivity 5–6 dS/m) at Sadhoke, Punjab, Pakistan, had lighter grain and higher Na content than control samples. Grains of three out of the four rices grown on saline soils had higher brown rice protein (higher nutritional value), less translucent grain, lower starch and amylose content, and lower K than their control samples, but these differences were not related to salinity tolerance. Alkali spreading value and gel consistency were not affected by culture in saline soil. Cooked rice Instron hardness increased in saline culture in two higher-protein samples of the four rices. Amylograph peak viscosity was suppressed by saline culture.
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  • 91
    ISSN: 1573-9104
    Keywords: rice ; grain quality ; inorganic and organic N fertilizers ; protein and lysine content ; season effect ; IR64
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract The effects of nitrogen fertilizer treatment and source (prilled urea, urea supergranule, fresh azolla, rice straw or sesbania or rice straw compost and their combinations) on grain quality were studied in the 1987 crops of variety IR64 at IRRI. Although fertilizer application improved grain yield, it improved protein content only in the case of urea supergranule, azolla and rice straw. Lysine contents of brown rice protein were similar in samples with no N fertilizer and those with the highest protein content in both seasons. Fertilizer treatment regardless of source tended to decrease weight and increase translucency of brown rice in both seasons. Effects on other grain properties were not consistent in both seasons. Season affected more grain properties than fertilizer treatment did, particularly translucency which was higher in the dry season than in the wet season.
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  • 92
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    Plant foods for human nutrition 40 (1990), S. 309-315 
    ISSN: 1573-9104
    Keywords: rice ; PAGE ; amino acid composition ; hydrophobicity index value
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Proteins and amino acids in four local rice (Oryza Sativa L.) varieties were identified. Albumin and globulin were extracted from rice seeds, and the major promoters of these proteins were investigated by polyacrylamide gel electrophoresis to show their patterns. Amino acid composition of the rice seed were determined quantitatively and qualitatively, and classified according to their acidic, basic and uncharged polar groups. Essential amino acids for each variety were determined, and the hydrophobicity index value of Amber 33 was (0.6078), Mishkhab 1 (0.63372), Hybrid 2 (0.6523) and Hwazawi (0.7411).
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  • 93
    ISSN: 1617-4623
    Keywords: Ty elements ; Saccharomyces cerevisiae ; Retrotransposon
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary To learn more about the variety of Ty elements capable of activating gene expression, we characterized 206 spontaneous Ty transpositions that activate the promoterless gene his3Δ4. Most of the Ty elements appear to be full-length, although a few deleted elements were recovered. Over 95% of the insertions belong to the Ty1 family, and the rest are Ty2 elements. The excessive number of Ty1 transpositions was unexpected because there are only 2-fold more Ty1 than Ty2 elements in the yeast strains used in the selection. However, there is 20-fold more Ty1 than Ty2 RNA present in these yeast strains. This difference in RNA level explains the greater number of Ty1 verses Ty2 transpositions at his3Δ4, because Ty elements transpose through an RNA intermediate. A similar association between the Ty transcript level and transpositional activation of his3Δ4 is obtained in cells expressing GAL1-promoted Ty2-H556 or Ty2-917 elements, but only if the element does not contain a marker. Genetically marked Ty2-H556NEO and-917NEO elements transpose into and activate his3Δ4 with the same efficiency as the previously characterized Ty1H3NEO element, but are underrepresented relative to the levels of TyNEO transcript. We also found that chromosomal Ty transcripts are even more abundant than previously estimated and comprise about 1% of total cellular RNA.
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  • 94
    ISSN: 1617-4623
    Keywords: Bacterial conjugation ; Cyanobacteria ; IncQ plasmids ; Saccharomyces cerevisiae ; DNA methylation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The promiscuous IncQ plasmid pKT210 (Cmr, Smr) is efficiently transferred by transpecific conjugation from Escherichia coli to the facultatively heterotrophic cyanobacterium Synechocystis PCC6803 when mobilized by a helper plasmid coding for IncP transfer functions. The IncQ plasmid is stably maintained in the cyanobacterium as an autonomously replicating multicopy plasmid with no detectable structural alterations and can be recovered by transformation back to E. coli when using a mcrA mcrB host. Thus, the replicative host-range of IncQ plasmids extends beyond purple bacteria to the distinct procaryotic taxon of cyanobacteria, allowing the use of these small plasmids as convenient cloning vectors in Synechocystis PCC6803 and presumably also in cyanobacteria that are not amenable to genetic transformation. In contrast, an IncQ plasmid bearing the TRP1 gene of Saccharomyces cerevisiae failed to replicate when transferred to that yeast by transformation.
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  • 95
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; AR07 gene ; Chorismate mutase ; GCN4 ; Transcriptional regulation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The gene AR07 encodes the monofunctional enzyme chorismate mutase, a branch point enzyme in the aromatic amino acid biosynthetic pathway in Saccharomyces cerevisiae. We investigated the transcription of the AR07 gene. Three 5′ ends at positions − 36, − 56 and − 73 and the 3′ end of the transcripts 146 bp downstream of the translational stop codon were mapped. As in the promoters of other aromatic amino acid biosynthetic genes, a recognition element for the GCN4 transcriptional activator of amino acid biosynthesis is located 425 base pairs (bp) upstream of the first transcriptional start point. This element binds GCN4 specifically in vitro. Northern analysis and determination of the specific enzyme activity reveals however, that the element is not sufficient to mediate transcriptional regulation by GCN4 in vivo. We thus suggest that in addition to a consensus sequence capable of binding the GCN4 protein other factors like, for example, chromatin structure, determine whether a recognition site for a transcription factor functions as an upstream activation sequence.
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  • 96
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; Splicing ; Nuclear structure
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We report on the characterization of the yeast prp20-1 mutant. In this temperature-sensitive mutant, multiple steps of mRNA metabolism are affected. The prp20-1 mutant strain showed alterations in mRNA steady-state levels, defective mRNA splicing and changes in transcription initiation or termination when shifted from the permissive to the non-permissive temperature. In addition, a change in the structure of the nucleus in these cells became apparent. Electron microscopy revealed an altered structure of the nucleoplasm of prp20-1 mutant cells when grown at the no-permissive temperature that was not observed in cells grown at the permissive temperature or in wild-type cells. The wild-type PRP20 gene was isolated and sequenced. The putative PRP20 protein has a molecular weight of 52 kDa. We found that the PRP20 gene is identical to the yeast SRM1 gene (Clark and Sprague 1989). In addition, the PRP20 protein sequence shows significant sequence similarity to the human RCC1 protein (Ohtsubo et al. 1987). This protein has been implicated in the control of chromosome condensation. Based on the phenotype of the prp20-1 mutant and the observed sequence similarity to the human RCC1 protein, we postulate that the yeast PRP20 protein is involved in the control of nuclear organization.
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  • 97
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    Molecular genetics and genomics 224 (1990), S. 111-118 
    ISSN: 1617-4623
    Keywords: Saccharomyces cerevisiae ; Translation ; Splicing ; Paromomycin
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The MSS51 gene product has been previously shown to be involved in the splicing of the mitochondrial pre-mRNA of cytochrome oxidase subunit I (COX1). We show here that it is specifically required for the translation of the COX1 mRNA. Furthermore, the paromomycin-resistance mutation (P inf454 supR ) which affects the 15 S mitoribosomal RNA, interferes, directly or indirectly, with the action of the MSS51 gene product. Possible roles of the MSS51 protein on the excision of COX1 introns are discussed.
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  • 98
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    Molecular genetics and genomics 224 (1990), S. 257-263 
    ISSN: 1617-4623
    Keywords: General Control ; Thermotolerance ; Saccharomyces cerevisiae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary In Saccharomyces cerevisiae starvation for a single amino acid activates the transcription of a set of genes belonging to different amino acid biosynthetic pathways (General Control, GC). We show that mutants affected in GC regulation are also affected in their response to thermal stress. Moreover, growth conditions that are known to induce heat shock proteins induce the GC response. However, unlike heat shock proteins, the transcriptional activator of GC, GCN4, is not induced after a short exposure to heat, and in gcn mutant strains induction of heat resistance is normal.
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  • 99
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    Molecular genetics and genomics 220 (1990), S. 269-276 
    ISSN: 1617-4623
    Keywords: GABA ; Saccharomyces cerevisiae ; Transcription ; DNA sequence ; Zinc finger
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The UGA3 gene of Saccharomyces cerevisiae is required for 4-aminobutyric acid (GABA)-dependent induction of the UGA1, UGA2 and UGA4 genes which encode the two GABA catabolic enzymes and a GABA-specific permease, respectively. Measurements of UGA1-specific transcripts show that induction of UGA1 correlates with accumulation of its RNA and requires a functional UGA3 gene. A 2 kb DNA fragment complementing the uga3 mutation was isolated and shown to contain the UGA3 gene. The primary structure of the UGA3 encoded protein was deduced from the DNA sequence, and contains an N-terminal, cysteine-rich motif similar in sequence to regions found in other fungal regulatory proteins and which are supposed to form zinc finger structures involved in DNA binding. Mutations were identified in the UGA3 genes isolated from uninducible and constitutive uga3 alleles. One case of intragenic complementation between two uninducible uga3 mutants is reported, indicating a possible oligomeric structure for UGA3. The role of UGA3 is discussed in relation to its genetic properties and its predicted structure.
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  • 100
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    Molecular genetics and genomics 220 (1990), S. 283-288 
    ISSN: 1617-4623
    Keywords: Chorismate mutase ; Saccharomyces cerevisiae ; Repression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The Saccharomyces cerevisiae ARO7 gene was cloned by screening a wild-type gene bank for complementation of an aro7 auxotrophic mutant. In vitro mutagenesis of the isolated plasmid (pJFB1) gave several transformants resistant to levels of the phenylalanine analogue 2-thienylalanine inhibitory to the wild-type transformant. Chorismate mutase assays indicated that two of the mutants (J14-26IV6 and J14-26IV9) were resistant to feedback inhibition by tyrosine displayed by wild-type strains. Analysis of the effect of other aromatic amino acids on chorismate mutase activity showed that tryptophan counteracted this inhibition. Analysis of the effect of tyrosine in the growth medium on enzyme activity indicated that the wild-type ARO7 gene was repressed by tyrosine, a phenomenon not previously reported. Two of the 2-thienylalanine resistant mutants (J14-26IV3 and J14-26IV9) appeared to be resistant to this repression. Transcriptional analysis confirmed that the level of ARO7 transcript decreased with increasing tyrosine concentration. In stain J14-26IV9 the ARO7 transcript level was not affected. J14-26IV9, therefore, appears to be a double mutant, resistant to both feedback inhibition and repression by tyrosine.
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