A Simple Method for Isolating DNA of High Yield and Quality from Cotton (shape Gossypium hirsutum L.) Leaves

An easy, reproducible and fast procedure to isolate DNA from cotton leaves is described. The addition of 0.5 M glucose in the extraction buffer avoids browning by polyphenolic compounds and improves the quality of DNA for molecular analysis. The DNA yield ranged between 150–400 mg per gram of fresh tissue. The DNA was suitable for digestion by restriction enzymes and amplificatiion by Taq DNA polymerase.     

Ligand-induced association of rat lymphocyte membrane proteins with the detergent-insoluble lymphocyte cytoskeletal matrix

There are two classes of membrane protein capping on the basis of ligand requirements. Surface immunoglobulin (Slg), the prototype of the first class, requires a single ligand for cap induction. RT1 (rat histocompatibility proteins) requires two antibodies for cap induction. The lateral mobility of Slg is relatively restricted compared with RT1. These differences may be due to differential interaction with the cytoskeleton. After ligand binding 71% of Slg becomes detergent insoluble and is associated with the lymphocyte cytoskeletal matrix. The insolubilization occurs at 4 degrees C and is not inhibited by sodium azide or cytoskeleton-active drugs. The insolubilized ligand-receptor complex can be solubilized by a cytoskeleton destabilizing buffer. In contrast, only 20% of RT1 becomes associated with the lymphocytic cytoskeleton after ligand binding. The ligand-induced receptor-cytoskeleton interaction influences capping behavior and may play a role in cell activation.     

Production of trace levels of carbon monoxide during methanogenesis on acetate and methanol

Summary Production of trace levels of carbon monoxide was consistently observed in the off-gas of a laboratory anaerobic digester fed Waste Activated Sludge. Inocula from this digester was enriched for acetate and methanol utilizing methanogenic populations. These enriched inocula were then monitored in batch assays for carbon monoxide and hydrogen production. Results demonstrated that carbon monoxide is produced during methanogenesis on both substrates. Subsequent utilization of CO was observed to occur after methane production was essentially complete for the assays conducted with methanol. Carbon monoxide evolution during methanogenesis on acetate displayed a markedly different trend from that observed from methanol.     

Production of Poly-β-Hydroxyalkanoic Acid by Pseudomonas cepacia

The possibility of using the nutritionally versatile bacterium Pseudomonas cepacia to produce poly-β-hydroxyalkanoic acid was evaluated. Chemostat culture showed that growth of P. cepacia became nitrogen limited when the molar carbon-to-nitrogen ratio of the medium fed into the fermentor was above 15. When grown under nitrogen limitation in batch culture with fructose as the sole source of carbon, P. cepacia accumulated poly-β-hydroxybutyric acid (PHB) in excess of 50% of the dry weight of its biomass. In batch culture, almost no PHB was produced until the onset of nitrogen limitation. After this point, PHB was produced at a linear rate of 0.12 g liter⁻¹ h⁻¹ (from a constant value of 1.6 g of cellular protein liter⁻¹). PHB produced by P. cepacia had a weight-average molecular weight of 5.37 × 10⁵ g mol⁻¹ and a polydispersivity index of 3.9. Poly(β-hydroxybutyric acid-β-hydroxyvaleric acid) copolymer was produced with a poly-β-hydroxybutyric acid-poly-β-hydroxyvaleric acid ratio of up to 30% by weight when propionic acid was added to the medium.     

Pathogenesis of murine cytomegalovirus infection in natural killer cell-depleted mice.

The effect of natural killer (NK) cells on the course of acute and persistent murine cytomegalovirus (MCMV) infection was examined by selectively depleting NK cell activity by inoculation of mice with antibody to asialo GM1, a neutral glycosphingolipid present at high concentrations on NK cells. The dose of MCMV required to cause 50% mortality or morbidity in control C57BL/6 mice dropped 4- and greater than 11-fold, respectively, in mice first treated with anti-asialo GM1. NK cell-depleted mice had higher (up to 1,000-fold) virus titers in their lungs, spleens, and livers at days 3, 5, 7, and 9 postinfection. Spleens and livers of control mice were virus-free by day 7 postinfection, and their lungs showed no signs of active infection at any time. In contrast, MCMV had disseminated to the lungs of NK cell-depleted mice by day 5, and these mice still had moderate levels of virus in their lungs, spleens, and livers at day 9. Markedly severe pathological changes were noted in the livers and spleens of NK cell-depleted, MCMV-infected mice. These included ballooning degeneration of hepatocytes and spleen necrosis. MCMV-infected, NK cell-depleted mice had severe spleen leukopenia, and their spleen leukocytes exhibited a significantly lower (up to 13-fold) response to the T cell mitogen concanavalin A when compared with those of uninfected and MCMV-infected controls. It appeared that NK cells exerted their most potent antiviral effect early in the infection, in a pattern correlating with interferon production and NK cell activation; treatment with anti-asialo GM1 later in infection had no effect on virus titers. The relative effect of NK cell depletion on MCMV pathogenesis depended on the injection route of the virus. NK cell depletion greatly augmented MCMV synthesis and pathogenesis in mice inoculated either intravenously or intraperitoneally but had no effect on the course of disease after intranasal inoculation, at any time point examined. One month after intraperitoneal inoculation of virus, NK cell depletion resulted in a six- to eightfold increase in salivary gland virus titers in persistently infected mice, suggesting that NK cells may be important in controlling virus synthesis in the salivary gland during persistent infection. This treatment did not, however, induce dissemination of virus to other organs. These data support the hypothesis that NK cells limit the severity, extent, and duration of acute MCMV infection and that they may also be involved in regulating the persistent infection.     

Immunohistopathological studies of blastomycosis: the use of labeled specific antigens in a hamster model of disease

We applied a modified immunofluorescence and immunoperoxidase method, utilizing labeled Blastomyces dermatitidis antigens, to look for specific antibody-bearing B/ plasma cells in the tissue infiltrates of blastomycosis lesions induced in hamsters. No specific anti-blastomyces antibodies were detectable by this method, although such antibodies were present in blood samples as demonstrated by routine immunodiffusion techniques. These studies suggest that humoral immune reactions do not play a major role in the pathogenesis of lesions of blastomycosis in hamsters.     

PROLIFERATION KINETICS OF THE MOUSE BLADDER AFTER IRRADIATION

The proliferative response of the mouse bladder was investigated, using continuous labelling with tritiated thymidine, at various times after a single dose of radiation. Bladder epithelial and vascular endothelial cells were studied. The cell turnover rate in unirradiated epithelium and endothelium was found to be extremely slow (in excess of 1 year). Irradiation with a single dose of 25 Gy resulted in compensatory proliferation of the epithelium but the response was not initiated for many months. At 3 months after irradiation there was little difference from the control proliferation rate, but from 6 to 22 months after irradiation (the end of the study) there was a period of sustained rapid proliferation with the cell turnover time reduced to approximately 1 week. The increase in proliferative activity observed at 22 months was found to be dose—dependent. Endothelial cells in the blood vessels of the submucosa also showed an increased turnover rate after irradiation and the timing of this reponse was found to be similar to that of the epithelium. The onset of compensatory proliferation in both cell types was found to coincide with marked histological and functional changes in the bladder. In this slowly proliferating tissue, the onset of rapid compensatory proliferation after irradiation is delayed and occurs at the time that functional impairment is observed. This supports the postulate that proliferation is unlikely to contribute much to the sparing effect of prolonged fractionated radiotherapy in slowly dividing tissues.     

Sensitivity of yeasts to lithium

A wide range of tolerance to Li⁺ has been found among 12 different yeasts. Concentrations that do not allow long-term growth also arrest growth of an actively growing culture within 2–5 h. At the same concentrations protein and RNA synthesis are inhibited with little or no lag period (<50 min) but respiration is not affected at these concentrations. Lower concentrations that do not inhibit growth, may impair sporulation. For given extracellular conditions, intracellular Li⁺ concentrations are lower in the more tolerant yeast strains.     

Molecular identification and dynamics of microbial communities in reactor treating organic household waste

The prokaryotic diversity associated with organic household waste (OHW), leachate (start-up inoculum), and mesophilic anaerobic digestion processes in the degradation of OHW for 44 and 90 days was investigated using a culture-independent approach. Bacterial and archaeal 16S rRNA and mcrA gene clone libraries were constructed from community DNA preparations. Bacterial clones were affiliated with 13 phyla, of which Firmicutes, Proteobacteria, and Bacteroidetes were represented in all libraries, whereas Actinobacteria, Thermotogae, Lentisphaerae, Acidobacteria, Chloroflexi, Cyanobacteria, Synergistetes, Spirochaetes, Deferribacteres, and Deinococcus-Thermus were exclusively identified in a single library. Within the Archaea domain, the Euryarchaeota phylum was the only one represented. Corresponding sequences were associated with the following orders of hydrogenotrophic methanogens: Methanomicrobiales (Methanoculleus genus) and Methanobacteriales (Methanosphaera and Methanobacterium genera). One archaeal clone was not affiliated with any order and may represent a novel taxon. Diversity indices showed greater diversity of Bacteria when compared to methanogenic Archaea.