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61.
A comparison of five methods for DNA isolation from liver and rumen flukes to perform ITS-2+ amplification 总被引:1,自引:0,他引:1
Five different DNA isolation methods (4 commercial kits and a modification of phenol-chloroform method) were compared for the discrimination of adults of Fasciola hepatica and Dicrocoelium dendriticum (liver flukes), and Calicophoron daubneyi (rumen fluke) collected from sheep in southern Italy. The second internal transcribed spacer (ITS-2) of ribosomal DNA (rDNA) plus flanking 5.8S and 28S sequence (ITS-2+) was amplified by polymerase chain reaction (PCR) from serial diluted DNA templates (6 ng - 60 fg) of each fluke species. Overall, in terms of efficiency in detection limit, the best results were obtained either with phenol-chloroform purification or with QIAamp DNA Mini Kit (Qiagen), but using this latter method, rapid, safe and not expensive, an increased level of sensitivity sufficient to detect small amounts of target-DNA was achieved. In addition, electrophoresis analysis following PCR also showed that ITS-2+ could be useful as a genetic marker for the molecular identification of F. hepatica, D. dendriticum and C. daubneyi in definitive and intermediate hosts. Furthermore, for the first time, the ITS-2 sequence of D. dendriticum was defined. 相似文献
62.
Properties and exploitation of oleosins 总被引:3,自引:0,他引:3
Oleosins stabilize oil bodies in seeds and other tissues and contain a unique hydrophobic domain which appears to be inserted into the oil matrix as an alpha-helical hairpin. The oleosin proteins may be exploited to stabilize emulsions while the ease of oil body preparation has led to the expression of bioactive proteins as oleosin fusions in molecular farming. 相似文献
63.
Mauricio A. Martins Damien C. Tully Michael A. Cruz Karen A. Power Marlon G. Veloso de Santana David J. Bean Colin B. Ogilvie Rujuta Gadgil Noemia S. Lima Diogo M. Magnani Keisuke Ejima David B. Allison Michael Piatak Jr. John D. Altman Christopher L. Parks Eva G. Rakasz Saverio Capuano III Ricardo Galler Myrna C. Bonaldo Jeffrey D. Lifson Todd M. Allen David I. Watkins 《Journal of virology》2015,89(21):10802-10820
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65.
Diaz-Perez, S. V., Crouch, V. W., and Orbach, M. J. 1996. Construction and characterization of a Magnaporthe grisea bacterial artificial chromosome library. Fungal Genet. Biol. 20, 280-288. A bacterial artificial chromosome (BAC) library of Magnaporthe grisea containing 4128 clones with an average insert size of 66-kb has been constructed. This library represents seven genome equivalents of M. grisea and has been demonstrated to be representative of the genome by screening for the presence of several single-copy genes and DNA markers. The utility of the library for use in map-based cloning projects was shown by the spanning of a nine-cosmid, 207-kb DNA contig with only 3 BAC clones. In addition, using a lys1-3 auxotroph, we have shown that BAC clones at least 113 kb can be transformed into M. grisea to screen for complementation of mutations. Thus, BACs isolated in chromosome walks can be rapidly screened for the presence of the sought after gene. The ease of construction of BAC libraries and of isolation and manipulation of BAC clones makes the BAC system an ideal one for physical analyses of fungal genomes. 相似文献
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67.
M. M. Rigano A. Romanelli A. Fulgione N. Nocerino N. D'Agostino C. Avitabile L. Frusciante A. Barone F. Capuano R. Capparelli 《Journal of peptide science》2012,18(12):755-762
Defensins are a class of cysteine‐rich proteins, which exert broad spectrum antimicrobial activity. In this work, we used a bioinformatic approach to identify putative defensins in the tomato genome. Fifteen proteins had a mature peptide that includes the well‐conserved tetradisulfide array. We selected a representative member of the tomato defensin family; we chemically synthesized its γ‐motif and tested its antimicrobial activity. Here, we demonstrate that the synthetic peptide exhibits potent antibacterial activity against Gram‐positive bacteria, such as Staphylococcus aureus A170, Staphylococcus epidermidis, and Listeria monocytogenes, and Gram‐negative bacteria, including Salmonella enterica serovar Paratyphi, Escherichia coli, and Helicobacter pylori. In addition, the synthetic peptide shows minimal (<5%) hemolytic activity and absence of cytotoxic effects against THP‐1 cells. Finally, SolyC exerts an anti‐inflammatory activity in vitro, as it downregulates the level of the proinflammatory cytokines TNF‐α and IFN‐γ. Copyright © 2012 European Peptide Society and John Wiley & Sons, Ltd. 相似文献
68.
Watanabe T Shinya K Watanabe S Imai M Hatta M Li C Wolter BF Neumann G Hanson A Ozawa M Yamada S Imai H Sakabe S Takano R Iwatsuki-Horimoto K Kiso M Ito M Fukuyama S Kawakami E Gorai T Simmons HA Schenkman D Brunner K Capuano SV Weinfurter JT Nishio W Maniwa Y Igarashi T Makino A Travanty EA Wang J Kilander A Dudman SG Suresh M Mason RJ Hungnes O Friedrich TC Kawaoka Y 《Journal of virology》2011,85(24):13195-13203
The first influenza pandemic of the 21st century was caused by novel H1N1 viruses that emerged in early 2009. An Asp-to-Gly change at position 222 of the receptor-binding protein hemagglutinin (HA) correlates with more-severe infections in humans. The amino acid at position 222 of HA contributes to receptor-binding specificity with Asp (typically found in human influenza viruses) and Gly (typically found in avian and classic H1N1 swine influenza viruses), conferring binding to human- and avian-type receptors, respectively. Here, we asked whether binding to avian-type receptors enhances influenza virus pathogenicity. We tested two 2009 pandemic H1N1 viruses possessing HA-222G (isolated from severe cases) and two viruses that possessed HA-222D. In glycan arrays, viruses possessing HA-222D preferentially bound to human-type receptors, while those encoding HA-222G bound to both avian- and human-type receptors. This difference in receptor binding correlated with efficient infection of viruses possessing HA-222G, compared to those possessing HA-222D, in human lung tissue, including alveolar type II pneumocytes, which express avian-type receptors. In a nonhuman primate model, infection with one of the viruses possessing HA-222G caused lung damage more severe than did infection with a virus encoding HA-222D, although these pathological differences were not observed for the other virus pair with either HA-222G or HA-222D. These data demonstrate that the acquisition of avian-type receptor-binding specificity may result in more-efficient infection of human alveolar type II pneumocytes and thus more-severe lung damage. Collectively, these findings suggest a new mechanism by which influenza viruses may become more pathogenic in mammals, including humans. 相似文献
69.
In the present work we have critically examined the use of the KCN-titration technique in the study of the control of the cellular respiration by cytochrome c oxidase (COX) in the presence of the mitochondrial membrane potential (Δψ(mito)) in HepG2 cells. We clearly show that the apparent high inhibition threshold of COX in the presence of maximal Δψ(mito) is due to the KCN-induced decrease of Δψ(mito) and not to a low control of COX on the mitochondrial respiration. The tight control exerted by COX on the Δψ(mito) provides further insights for understanding the pathogenetic mechanisms associated with mitochondrial defects in human neuromuscular degenerative disorders. 相似文献