Search results for "Fluorescent"
showing 10 items of 863 documents
Disassembly of structurally modified viral nanoparticles: characterization by fluorescence correlation spectroscopy.
2005
Abstract Analysis of the breakdown products of engineered viral particles can give useful information on the particle structure. We used various methods to breakdown both a recombinant enveloped virus and virus-like particles (VLPs) from two non-enveloped viruses and analysed the resulting subunits by fluorescence correlation spectroscopy (FCS). Analysis of the enveloped baculovirus, Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV), displaying the green fluorescent protein (GFP) fused to its envelope protein gp64 was performed in the presence and absence of 5 mM SDS and 25 mM DTT. Without treatment, the viral particle showed a diffusion time of 3.3 ms. In the presence of SDS…
Human cytomegalovirus US3 modulates destruction of MHC class I molecules
2012
Human cytomegalovirus (HCMV), a member of the Herpesviridae family, is proficient at establishing lifelong persistence within the host in part due to immune modulating genes that limit immune recognition. HCMV encodes at least five glycoproteins within its unique short (US) genomic region that interfere with MHC class I antigen presentation, thus hindering viral clearance by cytotoxic T lymphocytes (CTL). Specifically, US3 retains class I within the endoplasmic reticulum (ER), while US2 and US11 induce class I heavy chain destruction. A cooperative effect on class I down-regulation during stable expression of HCMV US2 and US3 has been established. To address the impact of US3 on US11-mediat…
DNA delivery to 'ex vivo' human liver segments.
2011
Hydrodynamic injection is an efficient procedure for liver gene therapy in rodents but with limited efficacy in large animals, using an 'in vivo' adapted regional hydrodynamic gene delivery system. We study the ability of this procedure to mediate gene delivery in human liver segments obtained by surgical resection. Watertight liver segments were retrogradely injected from hepatic vein with a saline solution containing a plasmid bearing the enhanced green fluorescent protein (eGFP) gene, under different conditions of flow rate (1, 10 and 20 ml s(-1)) and final perfused volume. Samples were cultured for 1 to 2 days and used for microscopy and molecular analysis of gene expression. The fluore…
Transgenic tools for proteomic analysis of ciliary transport
2015
Vision begins as photons are captured by photoreceptor cilia and light is converted into electrical signals that are then sent to the brain. As the photoreceptor cilium is not able to make its own proteins, all polypeptides needed for converting photons into electrical signals are synthetized in the cell body. How these molecules move from the cell body to cilia is still unclear. Opsin is one of the best-characterized transmembrane proteins. Our goal is to understand the mechanism of opsin transport into photoreceptor cilia. In this project, we use a combination of genetic and proteomic approaches in the zebrafish model. As the first step, we are constructing a transgenic line that expresse…
Human SPG11 cerebral organoids reveal cortical neurogenesis impairment
2018
Abstract Spastic paraplegia gene 11(SPG11)-linked hereditary spastic paraplegia is a complex monogenic neurodegenerative disease that in addition to spastic paraplegia is characterized by childhood onset cognitive impairment, thin corpus callosum and enlarged ventricles. We have previously shown impaired proliferation of SPG11 neural progenitor cells (NPCs). For the delineation of potential defect in SPG11 brain development we employ 2D culture systems and 3D human brain organoids derived from SPG11 patients’ iPSC and controls. We reveal that an increased rate of asymmetric divisions of NPCs leads to proliferation defect, causing premature neurogenesis. Correspondingly, SPG11 organoids appe…
Occurrence and genotypes of Giardia isolated from lambs in Spain
2009
Three hundred and eighty six faecal specimens were randomly collected from 1- to 3-month-old lambs from 16 farms in Spain to investigate the presence of different genotypes of Giardia duodenalis. Individual specimens were examined by IFA (Immunofluorescence assay) and beta-giardin PCR polymerase chain reaction. Cysts of G. duodenalis were shed by lambs in every flock analyzed, showing a prevalence by farms of 100%. The average prevalence of G. duodenalis for the 386 specimens was 42%, ranging from 8.3 to 80% depending on the farm. beta-giardin PCR positive samples were sequenced to determine the genotypes present at each farm and seven new subtypes of beta-giardin Assemblage E are reported …
The yeast osmosensitive mutant fps1Δ transformed by the cauliflower BobTIP1;1 aquaporin withstand a hypo-osmotic shock
2005
AbstractOsmoregulation plays an important role in cellular responses to osmotic stress in plants and in yeast. Aquaporins contribute to osmotic adjustment by facilitating transport of water or solutes across membranes. The tonoplastic water channel BobTIP1;1 (original name BobTIP26-1) genes are upregulated during dessication stress in cauliflower meristematic tissue. To investigate the physiological importance of BobTIP1;1, we expressed it in a Saccharomyces cerevisiae osmosensitive mutant fps1Δ. We showed that the defect in the yeast glycerol plasma membrane transporter is complemented by a plant cDNA encoding the aquaporin BobTIP1;1 which is localized in the vacuolar membrane of the compl…
Identification of Two Mannoproteins Released from Cell Walls of a Saccharomyces cerevisiae mnn1 mnn9 Double Mutant by Reducing Agents
1999
The cell wall of Saccharomyces cerevisiae represents some 30% of the total weight of the cell and is made up of β-glucans, mannose-containing glycoproteins (mannoproteins), and small amounts of chitin (9, 15). The mannoproteins can be divided into three groups according to the linkages that bind them to the structure of the cell wall: (i) noncovalently bound, (ii) covalently bound to the structural glucan, and (iii) disulfide bound to other proteins that are themselves covalently bound to the structural glucan of the cell wall (8). Our work has focused on the disulfide-bound mannoproteins, probably the least well known of the three groups mentioned above. Previous work (25) showed that trea…
A comparative study of the incorporation of a 1,6-beta-glucan and an O-glycosylated protein epitope into the cell wall of Candida albicans.
1996
The topological distribution of two epitopes in the cell wall of Candida albicans, the kinetics of their incorporation into the regenerating protoplast wall, and the effect of different antibiotics upon their incorporation and localization have been studied. To do so, two monoclonal antibodies that react against an O-glycosylated mannoprotein (1B12) and against a 1,6-beta-glucan epitope (JRR1) were used. The results show that the JRR1 epitope is localized in an internal layer of the cell wall, in contrast to the 1B12 epitope, which is superficial, and that the incorporation of the JRR1 epitope into walls of regenerating protoplasts precedes that of the 1B12 epitope. The JRR1 epitope is norm…
Development of a green fluorescent tagged strain of Aspergillus carbonarius to monitor fungal colonization in grapes.
2011
An enhanced green fluorescent protein has been used to tag an OTA-producing strain of Aspergillus carbonarius (W04-40) isolated from naturally infected grape berries. Transformation of the fungus was mediated by Agrobacterium tumefaciens. The most efficient transformation occurred when the co-cultivation was done with 104 conidia due to higher frequency of resistance colonies (894 per 104 conidia) and lower background obtained. To confirm the presence of the hph gene in hygromycin resistant colonies, 20 putative transformants were screened by PCR analysis. The hph gene was identified in all the transformants. Variation on the expression levels of the eGFP was detected among the transformant…