in vitro assay systems for gene mutation in mammalian cells
Báo cáo khoa học: Functional characterization of ecdysone receptor gene switches in mammalian cells docx
... partner for binding to ligand as well as for transactivation of reporter genes in cells An EcR gene switch that includes a fusion protein containing VP16 activation domain, GAL4 DNA binding domain ... EcRtvay) increased ligand sensitivity of EcR gene switch EcR gene switch in a monopartite format containing a fusion protein of VP16 activation domain, GAL4 DNA binding domain and the EcR DEF domains ... DNA binding domain and VP16 activation domain Various combinations of the EcR ligand binding domain, VP16 activation domain and GAL4 DNA binding domain were constructed (Fig 1A) and tested in NIH...
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Báo cáo khoa học: Identification of sodium salicylate as an hsp inducer using a simple screening system for stress response modulators in mammalian cells pptx
... examined the effects of SA on stress response in mammalian cells using a simple screening system, and revealed that SA is a potent Hsp inducer in mammalian cells, thereby protecting cells against ... into C3H10T1/2 cells (1 · 107 cells per 100 mm dish) with 30 lL lipofectAMINE reagent (Invitrogen) according to the manufacturer’s instructions, and incubated for 48 h Cells were then maintained ... HeLa cells were grown in Eagle’s Minimum Essential medium (Nissui Pharmaceutical) containing 10% bovine serum in a CO2 incubator at 37 °C Screening for Hsp modulators A reporter plasmid containing...
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Báo cáo y học: "An update on targeted gene repair in mammalian cells: methods and mechanisms" ppt
... demonstrating that cell cycle arrest in the S-phase occurs in ssODN-treated cells In these arrested cells cooperation between replication forks and the ssODN, including the search for homology, ... within episomal target genes in vitro as well as in vivo [4,8] SDF-mediated episomal gene repair has been reported in mouse embryonic stem cells and in human hematopoietic stem/progenitor cells ... determined by the amino-terminal end of the ZFs involved, and with the re-engineering of these domains, amino acid composition can be modified to induce highly specific ZFN-target binding [98] The central...
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Báo cáo khoa hoc:" CP-31398, a putative p53-stabilizing molecule tested in mammalian cells and in yeast for its effects on p53 transcriptional activity" potx
... lacking histidine and containing 20 mM 3-AT (figure 1C) In contrast, expression of three mutant http://www.jnrbm.com/content/3/1/5 forms of p53 [1] with point mutations in their DNA-binding domain ... stabilizing action of CP31398 in yeast cells and in human cells CP-31398, a compound isolated in an antibody-based in vitro screen, was reported to stabilize the p53 DNA-binding core domain and ... strain carrying an integrated bi-directional reporter gene construct in which a single p53 binding site from the human p21CIP1 promoter [12] was inserted between the divergent HIS3 and lacZ genes...
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Tài liệu Báo cáo khoa học: Extended half-life upon binding of destabilized intrabodies allows specific detection of antigen in mammalian cells pdf
... fused to d1-EGFP are maintained in the cells by binding to antigen Generation of soluble binding-defective and cysteine-deleted mutants To demonstrate that the observed intracellular colocalization ... of interest for rapidly selecting from a library reliable intrabodies in mammalian cells [5] In addition, the fact that only b-gal-expressing CHO cells were rescued from the cell mixture indicates ... Birmingham, UK) For the double staining, the cells were permeabilized in 0.1 m Tris ⁄ HCl pH 7.5 containing 0.2% (v ⁄ v) Triton X-100 for after fixation with paraformaldehyde and subsequently incubated...
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Báo cáo khoa học: Site-specific phosphorylation of MCM4 during the cell cycle in mammalian cells pot
... prevent binding of MCM complex to chromatin [12] In contrast, a recent finding suggests that an intermediate level of phosphorylation of MCM4 is required for chromatin binding during interphase ... cell cycle in a site-specific manner Cyclin-dependent protein kinase is involved in the phosphorylation of MCM4 To determine which kinase is involved in the phosphorylation of MCM4 in phases G2 ... MCM proteins on licensed chromatin in the G1 phase was detached from chromatin Phosphorylation of MCM4 and other factors may be involved in stronger binding of MCM4 protein to chromatin in the...
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Báo cáo Y học: Mydj2 as a potent partner of hsc70 in mammalian cells doc
... concentration, boiled for and analyzed by 10% SDS-mini PAGE and Western blotting Indirect immunofluorence CV1 cells growing in coverslips were incubated at 37 °C or 43 °C for h as indicated in the text ... were obtained by injecting a male rabbit with purified Mydj2 protein [30] Protein–protein interactions experiments For immunoprecipitations, cell lysates prepared in RIPA buffer were incubated ... NaCl/Pi and fixed for 10 min, at room temperature in 2% paraformaldehyde The cells were washed three times with cold NaCl/Pi and permeabilized by incubating in ice cold, absolute methanol for 3–5 at...
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Báo cáo khoa học: Dictyostelium differentiation-inducing factor-1 induces glucose transporter 1 translocation and promotes glucose uptake in mammalian cells pdf
... DIFs in mammalian cells [19] Yet, the mechanisms underlying the actions of DIFs in mammalian cells remain to be elucidated Meanwhile, in the course of a study of the in vitro actions of DIFs in mammalian ... supplemented with 75 lgÆmL)1 penicillin, 50 lgÆmL)1 streptomycin and 10% (v ⁄ v) fetal bovine serum RGM-1 cells were maintained in vitro at 37 °C (5% CO2) in DMEM containing a low concentration (1000 ... basal condition for the stimulation with insulin) Cells were then washed twice with Buffer A and incubated for 30 in the same buffer containing the reagents They were assayed for glucose uptake...
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Báo cáo Y học: Regulation of peptide-chain elongation in mammalian cells pptx
... since many of the phosphopeptides observed in response to insulin treatment in vivo are seen in maps generated from eEF1A phosphorylated by MS6K in vitro However, the in vivo maps also contain ... functional domains are also indicated (see key) as are the tryptophan (Trp85) required for calmodulin binding and the GxGxxG motif that may be involved in substrate binding (see text) within this motif ... may involve a glycine-rich motif, which would thus serve quite a different function in these kinases from the role of a similar motif in phosphate binding in other protein kinases The eEF2 kinase...
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Báo cáo khoa học: Expression studies of the core+1 protein of the hepatitis C virus 1a in mammalian cells pot
... data regarding this mechanism in cultured cells remain variable [15,21–24] In contrast, internal translation initiation has been identified only in mammalian cells, and recent evidence indicates ... that its detection in mammalian cells and in the HCV infectious system is elusive This study shows that intracellular levels of the core+1 protein in mammalian cells are strongly in uenced not only ... and 25 lm, respectively The protease inhibitor cocktail for mammalian extracts (containing AEBSF, aprotinin, leupeptin, bestatin, pepstatin a and E-64) was obtained from Sigma (St Louis, MO) Plasmid...
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Báo cáo y học: "Identification of secondary targets of N-containing bisphosphonates in mammalian cells via parallel competition analysis of the barcoded yeast deletion collection" ppsx
... MCF-7 cells in the S-phase (Figure S3a in Additional data file 1) In particular, the number of cycling cells in the S-phase increased from 16% to 21% for IBA, to 28% for RIS and to 38% for ALE ... compensated for by the presence of N-BPs, suggesting the occurrence of epistatic interactions involving the DBF4 gene Since DBF4 protein accumulates in the nuclei of G1-, S-, and M-phase-arrested cells ... electron microscope For immunogold labeling of tubulin, cells were fixed in neutral buffered 4% paraformaldehyde, dehydrated in graded ethanol and embedded in LRWhite resin Thin sections were collected...
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Trypsin-induced proteome alteration during cell subculture in mammalian cells potx
... trypsin-induced differential protein expression in cervical cancer cells It is interesting and important to know whether trypsineinduced protein alterations in MCF-7 cells are reproducible in other ... found in MCF-7 cells In contrast to an instantly up-regulated VDAC1 level was shown in MCF-7 cells, a significantly enhanced VDAC1 level in Hela cells starting from hr In summary, trypsinization-induced ... or membrane proteins Hence, this study describes a 2D-DIGE strategy to perform cellular proteins labeling for the monitoring of trypsin-induced proteome alterations in mammalian cells Materials...
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Báo cáo y học: "Autoprocessing of human immunodeficiency virus type 1 protease miniprecursor fusions in mammalian cells" pptx
... fusions are competent for autoprocessing in mammalian cells Darunavir and Indinavir inhibit fusion precursor autoprocessing in transfected 293T cells To further examine autoprocessing specificity, ... construct was engineered for mammalian expression to examine protease autoprocessing in transfected mammalian cells Autoprocessing of the fusion precursors carrying protease mutations that were ... autoprocessing as mutations blocking this cleavage have no significant influence on protease activity or Gag processing in transfected mammalian cells [19,20] In contrast, mutations blocking N-terminal cleavage...
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báo cáo khoa học: "Quantitative analysis of nanoparticle internalization in mammalian cells by high resolution X-ray microscopy" pps
... essentially yield 2D information Three-dimensional information can be obtained with TEM by continuous sectioning, but both the image taking and the image analysis are time consuming This is particularly ... required staining Figure shows that even without staining this imaging method can readily explore in detail the dissimilarities in the internalization of different AuNPs by different cell lines Specifically, ... more numerous in EMT cells, form larger cluster and are distributed more evenly in cytoplasm than in HeLa cells For comparison, we also show a similar image of naked AuNPs in CT-26 cells (Figure...
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Genetics of golgi apparatus regulation in mammalian cells
... screen in HeLa cells to identify molecular regulators Depletion of 159 signaling genes, nearly 20% of genes assayed, induced strong and varied perturbations in Golgi morphology Using bioinformatics ... and pancreatic β cells [35, 36] Interestingly, these tubules are only observed in mammalian cells, possibly an advanced feature added to cisternal stacking in mammalian evolution In other organisms ... in the lateral linking between stacks by tethering homotypic cisterna but in a non-redundant fashion, as single depletions resulted in ribbon unlinking [50, 51] While the role 13 of GRASP55 in...
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Molecular mechanisms underlying the regulation of the positioning and formation of the cleavage furrow in cytokinesis in mammalian cells
... action Inhibit myosin II ATPase activity by binding to myosin II-ADP-Pi Blebbistatin and inhibit the release of the ADP Inhibit actin polymerization by binding to the barb end of the FCytochalasin ... Overexpression an actin crosslinking protein α-actinin increased the accumulation of actin filaments at both equator and the entire cortex, leading to delayed cytokinesis and cytokinesis failure On ... effectors Inhibit aurora B kinase activity by competing with ATP to bind to Hesperadin the ATPase domain of aurora B Depolymerize microtubules by binding to tubulin dimmers and Nocodazole inhibiting...
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Expression studies of the proteolytic p10 fragment of the neuronal cdk5 activator in mammalian cells
... delivered into different mammalian cells including Cos-7, HEK293, PC12 and Neuro-2a and p10 fusion proteins were overexpressed in these cells The influences of p10 overexpression in these mammalian cells ... proteins such as actins and tubulins is too disruptive Instead, proteolysis is normally operated on the cytoskeleton cross-linking proteins such as filamin, fodrin or tau In addition, these proteins ... entering the cells and causing cell injury, cell medium was changed to anti-biotics free medium (500µl/well) before adding the FuGENE/DNA mixture into the wells (100µl/well) Finally, cells were incubated...
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Expression studies of the proteolytic p10 fragment of the neuronal cdk5 activator in mammalian cells
... delivered into different mammalian cells including Cos-7, HEK293, PC12 and Neuro-2a and p10 fusion proteins were overexpressed in these cells The influences of p10 overexpression in these mammalian cells ... proteins such as actins and tubulins is too disruptive Instead, proteolysis is normally operated on the cytoskeleton cross-linking proteins such as filamin, fodrin or tau In addition, these proteins ... entering the cells and causing cell injury, cell medium was changed to anti-biotics free medium (500µl/well) before adding the FuGENE/DNA mixture into the wells (100µl/well) Finally, cells were incubated...
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The effect of CDK1 mediated GOLGI vesiculation on mitotic progression in mammalian cells
... Prekeris et al 2001) During mitosis, the N-terminal domain of GM130, comprising the p115 binding site, is phosphorylated on serine 25 by CDK1/Cyclin B, thereby inhibiting p115 binding(Nakamura, Lowe ... (SOC) for 1hr at 37˚C with shaking at 250 revolutions per minute (rpm) in Unitron-Plus incubator shaker (Infors, Bottmingen, Switzerland), and incubated on Luria-Bertani Broth (LB) agar containing ... -200C Cells were then washed with PBS three times for minutes and permeabilized with 0.2% 17 Triton X-100 in PBS for 2-5 minutes After washing with PBS three times for min, cells were blocked for...
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gene delivery to mammalian cells, volume 1
... Loligomer MPG Galactosylated lysine Histidylated lysine, HK (LARL)6 Domains DNA binding, NLS DNA binding, NLS DNA binding, galactose DNA binding pH-buffering DNA binding, hydrophobic Optimal peptide/ ... Maintain MDA-MB-435, MDA-MB-435, and CHO, NHDF, and BAE (for gene expression) cells in DMEM containing 10% FCS and mM glutamine Perform experiments on NHDF between passages and Maintain BAE cells, ... Reporter Assay Kit Intergen 800-431-4505 www.intergen.com Invitrogen 800-955-6288 www.invitrogen.com InvivoGen 888-457-5873 www.invivogen.com Marker Gene Tech Inc 541-342-3760 www.markergene.com...
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- incorporating geographic information systems for business in higher education
- chitosan conjugated dna nanoparticle delivery systems for gene therapy
- selection counter selection for gene mutation replacement and fusion
- coevolution in gene for gene systems
- generation of fluorescent igg fusion proteins in mammalian cells
- rnai screening in mammalian cells to identify novel host cell molecules involved in the regulation of viral infections
- assay systems for addressing new targets
- measuring phosphorylation specific changes in response to kinase inhibitors in mammalian cells using quantitative proteomics
- regulation of iron metabolism in mammalian cells
- in cell nmr in mammalian cells part 1
- in cell nmr in mammalian cells part 2
- in cell nmr in mammalian cells part 3
- macroautophagy in mammalian cells
- reconstituted fusion liposomes for gene transfer in vitro and in vivo
- assay systems for chromosome damage
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