Isolation and characterization of microsatellite loci in the woolly mouse opossum, Micoureus paraguayanus (Marsupialia : Didelphimorphia)

Five microsatellite loci were isolated and characterized within the woolly mouse opossum (Micoureus paraguayanus), a Neotropical marsupial, using an enrichment cloning procedure. Between four and seven alleles were detected per locus, with expected heterozygosity ranging from 0.358 to 0.560. These microsatellites should provide useful markers in a variety of genetic analyses to examine parentage, inbreeding, population structure and population dynamics in fragmented forest habitats.

Trypanosoma cruzi: identification and characterization of a novel ribosomal protein L27 (TcrL27) that cross-reacts with an affinity-purified anti-Sm antibody

Small nuclear ribonucleoproteins (snRNPs)are involved in trans-splicing processing of pre-mRNA in Trypanosoma cruzi. To clone T. cruzi snRNPs we screened an epimastigote cDNA library with a purified antibody raised against the Sm-binding site of a yeast sequence. A clone was obtained containing a 507 bp-insert with an ORF of 399 bp and coding for a protein of 133 amino acids. Sequence analysis revealed high identity with the L27 ribosomal proteins from different species including: Canis familiaris, Homo sapiens, Schizosaccharomyces pombe and Saccharomyces cerevisiae. This protein has not been previously described in the literature and seems to be a new ribosomal protein in T. cruzi and was given the code TcrL27. To express this recombinant T. cruzi L27 ribosomal protein in E. coli, the insert was subcloned into the pET32a vector and a 26 kDa recombinant protein was purified. Immunoblotting studies demonstrated that this purified recombinant protein was recognized by the same anti-Sm serum used in the library screening as well as by chagasic and systemic lupus erythemathosus (SLE) sera. Our results suggest that the T. cruzi L27 ribosomal protein may be involved in autoimmunity of Chagas disease.

Development and characterization of an equine behaviour chamber and the effects of amitraz and detomidine on spontaneous locomotor activity

This report describes the development of a behaviour chamber and the validation of the chamber to measure locomotor activity of a horse, Locomotor activity was detected by four Mini-beam sensors and recorded on a data logger every 5 min for 22 h. Horses were more active during daytime than in the evening, which was at least partially related to human activity in their surroundings. To validate the ability of the chambers to detect changes in activity, fentanyl citrate and xylazine HCl, agents well-characterized as a stimulant and a depressant, respectively, were administered to five horses. Fentanyl citrate (0.016 mg/kg) significantly increased locomotor activity which persisted for 30 min, Xylazine HCl (1 mg/kg) significantly reduced locomotor activity for 90 min. Amitraz produced a dose-dependent decrease in locomotor activity, lasting 75 min for the 0.05 mg/kg dose, 120 min for the 0.10 mg/kg dose, and 180 min for the 0.15 mg/kg dose, In a separate experiment, yohimbine administration immediately reversed the sedative effect of amitraz, This suggests there is a similarity in the mode of action of amitraz, xylazine and detomidine, as yohimbine acts primarily by blocking central alpha 2-adrenoceptors that are stimulated by agents like xylazine, There was also a significant decrease in locomotor activity following injection of detomidine (0.02, 0.04 and 0.08 mg/kg) for 1.5, 3.5 and 5.0 h, respectively, the locomotor chamber is a useful, sensitive and highly reproducible tool for measuring spontaneous locomotor activity in the horse, which allows investigators to determine an agent's average time of onset, duration and intensity of effect on movement.

Partial purification and characterization of pectin methylesterase from acerola (Malpighia glabra L.)

The enzyme pectin methylesterase (PME) is present in acerola fruit and was partially purified by gel filtration on Sephadex G-100. The results of gel filtration showed different PME isoforms. The total PME (precipitated by 70% salt saturation) and one of these isoforms (fraction from Sephadex G-100 elution) that showed a molecular mass of 15.5 +/- 1.0 kDa were studied. The optimum pH values of both forms were 9.0. The total and the partially purified PME showed that PME specific activity increases with temperature, the total acerola PME retained 13.5% of its specific activity after 90 min of incubation at 98 degreesC. The partially purified acerola (PME isoform) showed 125.5% of its specific activity after 90 min of incubation at 98 degreesC. The K-m values of the total PME and the partially purified PME isoform were 0.081 and 0.12 mg/mL, respectively. The V-max values of the total PME and the partially purified PME were 2.92 and 6.21 mumol/min/mL/mg of protein, respectively.

Identification and characterization of different accessions of itchgrass (Rottboellia cochinchinensis)

Studies were conducted to identify and characterize different accessions of itchgrass. Seeds were collected in the counties of Aramina, Campinas, Dumont, Igarapava, Jaboticabal, and Ribeirao Preto, all in the state of São Paulo, Brazil. Accessions were characterized based on dimensions of their stomata, stomatal index (SI), and length and width of their seed (caryopses and husk). Chromosome number and length also were determined, and accessions were further differentiated using molecular markers (polymerase chain reaction [PCR]). Itchgrass from Ribeirao Preto had much longer and narrower seeds than those from the other locations, and their husks were longer as well. Accessions had similar SIs, both on the abaxial and adaxial leaf surfaces. Stomata from Campinas and Igarapava accessions were longer and wider, whereas those from Dumont and Ribeirao Preto were similar and smaller than all others. The accession from Ribeirao Preto is diploid (2n = 20); the rest are polyploid, with the total length of chromosomes smaller than all others. These differences were confirmed by molecular differentiation (PCR).

Purification and characterization of polygalacturonase produced by thermophilic Thermoascus aurantiacus CBMAI-756 in submerged fermentation

An extracellular polygalacturonase was isolated from 5-day culture filtrates of Thermoascus aurantiacus CBMAI-756 and purified by gel filtration and ion-exchange chromatography. The enzyme was maximally active at pH 5.5 and 60-65 degrees C. The apparent K (m) with citrus pectin was 1.46 mg/ml and the V (max) was 2433.3 mu mol/min/mg. The apparent molecular weight of the enzyme was 30 kDa. The enzyme was 100% stable at 50 degrees C for 1 h and showed a half-life of 10 min at 60 degrees C. Polygalacturonase was stable at pH 5.0-5.5 and maintained 33% of initial activity at pH 9.0. Metal ions, such as Zn+2, Mn+2, and Hg+2, inhibited 50, 75 and 100% of enzyme activity. The purified polygalacturonase was shown to be an endo/exo-enzyme, releasing mono, di and tri-galacturonic acids within 10 min of hydrolysis.

PROCESSING AND CHARACTERIZATION OF LOW-VOLTAGE VARISTORS

ZnO seed particles and Cr2O3 were used in this study to control the microstructure of ZnO varistors. The seed particles were prepared by adding 1.0 mol % BaO to ZnO. The powder was then calcined at 800-degrees-C for 2 h, pressed into pellets and sintered at 1400-degrees-C for 8 h. The sintered ZnO was ground and the BaO eliminated by washing in water. The remaining ZnO powder was classified into a size fraction ranging from 38 to 149 mum. The addition of a small amount (1 weight %) ZnO seed grains produces varistors with low breakdown voltages (7.6 V/mm) and an alpha coefficient of approximately 10. The addition of Cr2O3 stabilizes the spinel phase yielding a more homogeneous microstructure, but degraded electrical behaviour of the ZnO varistor.

Production and characterization of alpha-amylase from Rhizopus sp

A strain of Rhizopus sp. screened among more than 800 filamentous fungi showed great ability to produce a thermostable alpha-amylase by solid state fermentation. The best production was obtained with a bran moisture content of 40% when the enzyme activity reached 60 EU/g. of medium. During the purification procedures, a column of DEAE-Sephadex A-50 separated the enzyme in two fractions and the larger (85% of the total activity) showed optimum pH in a range from 4.0 to 5.6. Optimum temperature was found at 60-65 degrees C and in this range no loss of activity was observed after 60 min. of treatment in pH 5,0. Its K-m and V-m are, respectively, of 5.0 mg/ml of starch and 10,01 uMol of reducing sugar/min./mg. of protein. Its molecular weight was calculated in 64.000 by gel filtration in Sephadex G-200. The dextrinization power of the enzyme was observed preferentialy on substrates compound by chains with higher ramifications, that is: amylopectin > starch > amylose. Other aspects of the enzyme pattern action are also discussed.

Identification and characterization of Colletotrichum spp. affecting fruit after harvest in Brazil

Colletotrichum spp. cause anthracnose in various fruits post-harvest and are a particularly important problem in tropical and subtropical fruits. The disease in fruits of avocado, guava, papaya, mango and passion fruit has been reported to be caused by C. gloeosporioides, and in banana by C. musae. In subtropical and temperate crops such apple, grape, peach and kiwi, the disease is caused by C. acutatum. The variation in pathogenic, morphological, cultural and molecular characteristics of Brazilian isolates of Colletotrichum acutatum Simmonds and isolates from post-harvest decays of avocado, banana, guava, papaya, mango and passion fruit was evaluated. The fruits were inoculated with mycelium of C. acutatum, Colletotrichum spp. and C. musae on a disc of potato dextrose agar. The morphological, cultural and molecular characteristics studied were conidia morphology, colony growth at different temperatures, colony coloration and PCR with primers CaInt2 and ITS4 for C. acutatum and CgInt and ITS4 for C. gloeosporioides. C. acutatum was pathogenic to avocado, guava, papaya, mango and passion fruit, but it was not pathogenic to banana. The morphological, cultural and molecular studies indicated that the avocado, papaya, mango and passion fruit isolates were C. gloeosporioides. The natural guava isolate was identified as C. acutatum, which had not been found previously to produce anthracnose symptoms on guava in Brazil.

Synthesis and structure of a new heteronuclear (eta(6)-arene) tricarbonylchromium compound incorporating propargyl amine unit

Reaction of equimolar amounts of [Co-2(CO)(6)(dppm)] and [Cr(eta(6)-C6H5)(C=-CCH2NMe2)(CO)(3)] (1) in benzene gives the novel heterotrimetallic complex [Cr(eta(6)-C6H5){(mu(2)-eta(2)-C=CCH2NMe2)Co-2(CO)(4)(dppm)}(CO)(3)] (2) in 40% yield. The X-ray structure analysis of 2 reveals that the Cr(CO)(3) tripod adopts a nearly anti-eclipsed conformation. (C) 2002 Elsevier B.V. B.V. All rights reserved.

Purification and characterization of beta-fructosidase with inulinase activity from Aspergillus niger-245

Aspergillus niger - 245 a strain isolated from soil samples showed good beta -fructosidase activity when inoculated in medium formulated with dahlia extract tubers. The enzyme was purified by precipitation in ammonium sulphate and percolated in DEAE-Sephadex A-50 and CM-cellulose columns, witch showed a single peack in all the purification steps, maintaining the I/S ratio between 0.32 to, 0.39. Optimum pH for inulinase activity (I) was between 4.0 - 4.5 and for invertase activity (S) between 2.5 and 50. The optimum temperature was 60 degrees .C for both activities and no loss in activity was observed when it was maintained at this temperature for 30 min. The K-m value was 1.44 and 5.0 respectively, for I and S and V-m value 10.48 and 30.55 respectively. The I activity was strongly inhibited by Hg2+ and Ag+ and 2 x 10(-3) M of glucose, but not by fructose at the same concentration. The enzyme showed an exo-action mechanism acting on the inulin of different origins. In assay conditions total hydrolysis of all the frutans was obtained although it has shown larger activity on the chicory inulin than that one from artichoke Jerusalem and dahlia, in the first 30 min. The obtained results suggested that the enzyme presented good potential for industrial application in the preparing the fructose syrups.