control gfp expression vector Search Results


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  • 92
    Thermo Fisher gfp expression control vector
    Gfp Expression Control Vector, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Shanghai Genechem control vector expressing gfp
    Expression of <t>GFP</t> and Foxc2 genes after <t>transfection.</t> (A) Transfection efficiency at various MOIs (10, 50, 100 and 200) 72 hours after transfection (×100). (B) Cell proliferation after transfection in the Lv-Foxc2 and Lv-GFP groups detected using MTT assay. (C) Foxc2 gene expression measured by real-time PCR 72 hours after transfection. GAPDH was used as a housekeeping gene. (D) Foxc2 protein expression evaluated by western blot 72 hours after transfection. Results were expressed as mean±SD of triplicate experiments. #p
    Control Vector Expressing Gfp, supplied by Shanghai Genechem, used in various techniques. Bioz Stars score: 85/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Amaxa expression vector pmax gfp
    Expression of <t>GFP</t> and Foxc2 genes after <t>transfection.</t> (A) Transfection efficiency at various MOIs (10, 50, 100 and 200) 72 hours after transfection (×100). (B) Cell proliferation after transfection in the Lv-Foxc2 and Lv-GFP groups detected using MTT assay. (C) Foxc2 gene expression measured by real-time PCR 72 hours after transfection. GAPDH was used as a housekeeping gene. (D) Foxc2 protein expression evaluated by western blot 72 hours after transfection. Results were expressed as mean±SD of triplicate experiments. #p
    Expression Vector Pmax Gfp, supplied by Amaxa, used in various techniques. Bioz Stars score: 85/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    5 PRIME gfp control vector
    Western Blot of WT <t>GFP</t> and GFP Y39TAG mutant expression in a cell-free translation system. Synthesis of WT GFP and the GFP Y39TAG mutant was performed using the <t>RTS</t> E. coli HY Kit, to which the corresponding plasmid (500 µg/mL), purified Mj TyrRS and cognate suppressor Mj tRNA CUA (tRNA) or T-stem modified tRNA CUA Opt (denoted as *) were added. (A) Expression of WT GFP and the GFP Y39TAG mutant in the presence of Mj TyrRS (300 µg/mL) and synthetic Mj tRNA CUA (60 µg/mL). The band at 28 kDa corresponds to full-length GFP. (B) Western blot analysis demonstrates enhanced GFP Y39TAG protein expression as a function of increased Mj TyrRS concentrations in a cell-free reaction medium supplied with Mj tRNA CUA (60 µg/mL – top panel and 450 µg/mL – bottom panel). ( C ) Dependence of GFP Y39TAG yield on the type and concentration of nonsense suppressor, as visualized by Western blot.
    Gfp Control Vector, supplied by 5 PRIME, used in various techniques. Bioz Stars score: 88/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    TaKaRa control vector expressing gfp
    Western Blot of WT <t>GFP</t> and GFP Y39TAG mutant expression in a cell-free translation system. Synthesis of WT GFP and the GFP Y39TAG mutant was performed using the <t>RTS</t> E. coli HY Kit, to which the corresponding plasmid (500 µg/mL), purified Mj TyrRS and cognate suppressor Mj tRNA CUA (tRNA) or T-stem modified tRNA CUA Opt (denoted as *) were added. (A) Expression of WT GFP and the GFP Y39TAG mutant in the presence of Mj TyrRS (300 µg/mL) and synthetic Mj tRNA CUA (60 µg/mL). The band at 28 kDa corresponds to full-length GFP. (B) Western blot analysis demonstrates enhanced GFP Y39TAG protein expression as a function of increased Mj TyrRS concentrations in a cell-free reaction medium supplied with Mj tRNA CUA (60 µg/mL – top panel and 450 µg/mL – bottom panel). ( C ) Dependence of GFP Y39TAG yield on the type and concentration of nonsense suppressor, as visualized by Western blot.
    Control Vector Expressing Gfp, supplied by TaKaRa, used in various techniques. Bioz Stars score: 91/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    80
    TaKaRa fluorescence protein gfp expression vector pd2egfp control
    Western Blot of WT <t>GFP</t> and GFP Y39TAG mutant expression in a cell-free translation system. Synthesis of WT GFP and the GFP Y39TAG mutant was performed using the <t>RTS</t> E. coli HY Kit, to which the corresponding plasmid (500 µg/mL), purified Mj TyrRS and cognate suppressor Mj tRNA CUA (tRNA) or T-stem modified tRNA CUA Opt (denoted as *) were added. (A) Expression of WT GFP and the GFP Y39TAG mutant in the presence of Mj TyrRS (300 µg/mL) and synthetic Mj tRNA CUA (60 µg/mL). The band at 28 kDa corresponds to full-length GFP. (B) Western blot analysis demonstrates enhanced GFP Y39TAG protein expression as a function of increased Mj TyrRS concentrations in a cell-free reaction medium supplied with Mj tRNA CUA (60 µg/mL – top panel and 450 µg/mL – bottom panel). ( C ) Dependence of GFP Y39TAG yield on the type and concentration of nonsense suppressor, as visualized by Western blot.
    Fluorescence Protein Gfp Expression Vector Pd2egfp Control, supplied by TaKaRa, used in various techniques. Bioz Stars score: 80/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    System Biosciences Inc control vector pmirna1 gfp
    a A luciferase reporter expression assay was performed to examine whether miR-520d-5p actually targets TEAD1 and GATAD2B . Out of the more than ten predicted binding sites in the 3′UTRs of TEAD1 and GATAD2B , two sequences [TTACAAG and TACAAAG in the 3′UTR of TEAD1 ( top ) or GATAD2B ( bottom )] were chosen to test for miR-520d-5p binding. These sites were predicted based on the four databases described above. b A luciferase reporter expression assay revealed that miR-520d-5p bound to both the 3′UTR of TEAD1 and the 3′UTR of GATAD2B . In contrast, a mismatch of miR-520d-5p (a control for standardizing data) and a scramble group did not significantly bind to the 3′UTR sites, resulting in a failure to induce luciferase expression. Using a luciferase reporter expression assay, the potent sites of miR-520d-5p binding to the 3′UTR were identified and representatively shown at base pairs 4,569–4,581 and 7,463–7,776 in the 3′UTR of TEAD1 and 3,622–3,639 in the 3′UTR of GATAD2B , respectively. The minus signs (−) in the synthesized miR-520d-5p sequence represent the mismatches in the synthesized miR-520d-3p, and the minus signs in the control vector <t>pMIRNA1/GFP</t> sequence represent the mismatch sequences within miR-520d-5p that was expressed from pMIRNA1-520d-5p/GFP ( n = 4). The data were analyzed by t test (* P
    Control Vector Pmirna1 Gfp, supplied by System Biosciences Inc, used in various techniques. Bioz Stars score: 86/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Addgene inc gfp only control vector
    dCas9 3A3L or dCas9 3A targeting prevents senescence and increases proliferation. a , b Proliferation was assessed using a colorimetric assay 10, 15 and 20 days after targeting dCas9 3A3L to HIC1 , RASSF1 , PTEN and CDKN2A and magnetic sorting in primary myoepithelial cells from donors 1, 2 and 3. Dotted line represents the average absorbance from three dCas9 3A3LΔ targeted wells at each timepoint. The graphs show the average difference in absorbance as fold change compared to 3A3LΔ average (mean ± SEM, n = 3). c Cumulative population doublings over time for untransfected primary myoepithelial cells and those transfected with dCas9 3A3L or 3A3L∆ targeting the four genes without magnetic sorting from donor one. Fifty thousand cells were seeded at the start of each passage and cells counted after 5 days (mean ± SEM, n = 3, where error bars are smaller than points plotted they are not shown). ( d ) Light microscopy images of β-gal staining from untransfected (top left) primary myoepithelial cells from donor 1 and those transfected with dCas9 3A3L (bottom two) or 3A3L∆ (top right) targeting the four genes without magnetic sorting 20 days after transfection, blue/green colouring shows β-gal +ve cells; scale bar represents 125 µm. e Proliferation was assessed using a colorimetric assay 15, 20 and 25 days after targeting the four genes with dCas9 3A3L (red), dCas9 3A (orange), dCas9 3A-C706A-3L (yellow), dCas9 3A3L∆ (green) or dCas9m4 (pale green), or targeting only dCas9 3A3L without gRNAs (blue) without magnetic sorting using donor 1 myoepithelial cells. The data is shown as fold change compared to the average absorbance from 3 wells without cells (mean ± SEM, n = 3). f The percentage of <t>GFP</t> +ve cells after infection with no virus (green), <t>hTERT</t> with GFP marker (pink) or GFP control (purple). At infection day 0 cells are 38 days post-transfection with dCas9 3A3L targeting the four genes (mean ± SEM; n = 3, error bars are not shown where smaller than points plotted)
    Gfp Only Control Vector, supplied by Addgene inc, used in various techniques. Bioz Stars score: 91/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Vector Biolabs raav gfp control vector
    VGF mRNA and eGFP expression patterns are similar following <t>rAAV-VGF</t> infusio n into the hypothalamus of both Siberian hamsters and mice . Representative pictures of eGFP expression (left) and VGF mRNA expression via in situ hybridisation (right) in the hypothalamus of Siberian hamsters (A and B) and mice (C and D) receiving bilateral injections of either <t>rAAV-GFP</t> (A and C) or rAAV-VGF (B and D) directed to the PVN. Scale bar = 2.5 mm.
    Raav Gfp Control Vector, supplied by Vector Biolabs, used in various techniques. Bioz Stars score: 88/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Genechem adcmv gfp control vector
    VGF mRNA and eGFP expression patterns are similar following <t>rAAV-VGF</t> infusio n into the hypothalamus of both Siberian hamsters and mice . Representative pictures of eGFP expression (left) and VGF mRNA expression via in situ hybridisation (right) in the hypothalamus of Siberian hamsters (A and B) and mice (C and D) receiving bilateral injections of either <t>rAAV-GFP</t> (A and C) or rAAV-VGF (B and D) directed to the PVN. Scale bar = 2.5 mm.
    Adcmv Gfp Control Vector, supplied by Genechem, used in various techniques. Bioz Stars score: 88/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Genecopoeia gfp expressing lentiviral control vector
    VGF mRNA and eGFP expression patterns are similar following <t>rAAV-VGF</t> infusio n into the hypothalamus of both Siberian hamsters and mice . Representative pictures of eGFP expression (left) and VGF mRNA expression via in situ hybridisation (right) in the hypothalamus of Siberian hamsters (A and B) and mice (C and D) receiving bilateral injections of either <t>rAAV-GFP</t> (A and C) or rAAV-VGF (B and D) directed to the PVN. Scale bar = 2.5 mm.
    Gfp Expressing Lentiviral Control Vector, supplied by Genecopoeia, used in various techniques. Bioz Stars score: 90/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa control vector expressing gfp protein
    Decreased dendritic complexity in <t>DSCAM-overexpressing</t> hippocampal neurons. A , Representative micrographs of hippocampal neurons transfected at DIV7 with the indicated constructs. Neuronal morphology was visualized by <t>GFP</t> immunocytochemistry 2 d after transfection. Dendrites were identified as MAP2-positive neurites. Scale bars, 60 μm. B , Sholl analysis of the neurons transfected with GFP or DSCAM-IRES-GFP vectors. C , Total dendritic length of neurons transfected with the indicated constructs. D , Number of primary dendrites of neurons transfected with the indicated constructs. E , Somatic area of neurons transfected with the indicated plasmids. Mean values are shown (12–15 neurons); the error bars indicate the SEM, and asterisks denote statistically significant differences: * p
    Control Vector Expressing Gfp Protein, supplied by TaKaRa, used in various techniques. Bioz Stars score: 85/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Lonza gfp vector
    <t>BATF</t> silencing enhanced cytokine secretions by PPD-specific T cells in PBMCs in patients with ATB. (A) Representative fluorescence microscopy image of transfected primary human PBMCs after 24 h of electroporation with 2 μg <t>GFP</t> Vector, and red arrow indicated transfected PBMCs containing GFP (× 200). (B) Transfection efficiency of primary human PBMCs 24 h post electroporation were assessed by analysis of GFP protein expression by flow cytometry. (C) Silencing of BATF by a BATF siRNA SMART pool in primary human PBMCs from ATB patients measured by real-time quantitative PCR. Expression normalized to a housekeeping gene ( GAPDH ) is presented as fold change relative to negative control siRNA. (D) PBMCs were electroporated with indicated siRNA, then cultured with M. tb -specific antigen PPD (25 μg/mL) overnight, and IL-2 or IFN-γ mRNA levels were measured by real-time quantitative PCR. The information of controls and cell viability after transfection was provided in the Methods section BATF Small Interfering RNA (siRNA) Knockdown in ATB Patients ( 11 ). Data are expressed as mean ± SEM ( n = 7). * P
    Gfp Vector, supplied by Lonza, used in various techniques. Bioz Stars score: 91/100, based on 34 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    GenePharma Company control vector pglv3 h1 gfp puro
    <t>BATF</t> silencing enhanced cytokine secretions by PPD-specific T cells in PBMCs in patients with ATB. (A) Representative fluorescence microscopy image of transfected primary human PBMCs after 24 h of electroporation with 2 μg <t>GFP</t> Vector, and red arrow indicated transfected PBMCs containing GFP (× 200). (B) Transfection efficiency of primary human PBMCs 24 h post electroporation were assessed by analysis of GFP protein expression by flow cytometry. (C) Silencing of BATF by a BATF siRNA SMART pool in primary human PBMCs from ATB patients measured by real-time quantitative PCR. Expression normalized to a housekeeping gene ( GAPDH ) is presented as fold change relative to negative control siRNA. (D) PBMCs were electroporated with indicated siRNA, then cultured with M. tb -specific antigen PPD (25 μg/mL) overnight, and IL-2 or IFN-γ mRNA levels were measured by real-time quantitative PCR. The information of controls and cell viability after transfection was provided in the Methods section BATF Small Interfering RNA (siRNA) Knockdown in ATB Patients ( 11 ). Data are expressed as mean ± SEM ( n = 7). * P
    Control Vector Pglv3 H1 Gfp Puro, supplied by GenePharma Company, used in various techniques. Bioz Stars score: 89/100, based on 48 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Vector Biolabs control gfp expressing adenovirus adgfp
    <t>BATF</t> silencing enhanced cytokine secretions by PPD-specific T cells in PBMCs in patients with ATB. (A) Representative fluorescence microscopy image of transfected primary human PBMCs after 24 h of electroporation with 2 μg <t>GFP</t> Vector, and red arrow indicated transfected PBMCs containing GFP (× 200). (B) Transfection efficiency of primary human PBMCs 24 h post electroporation were assessed by analysis of GFP protein expression by flow cytometry. (C) Silencing of BATF by a BATF siRNA SMART pool in primary human PBMCs from ATB patients measured by real-time quantitative PCR. Expression normalized to a housekeeping gene ( GAPDH ) is presented as fold change relative to negative control siRNA. (D) PBMCs were electroporated with indicated siRNA, then cultured with M. tb -specific antigen PPD (25 μg/mL) overnight, and IL-2 or IFN-γ mRNA levels were measured by real-time quantitative PCR. The information of controls and cell viability after transfection was provided in the Methods section BATF Small Interfering RNA (siRNA) Knockdown in ATB Patients ( 11 ). Data are expressed as mean ± SEM ( n = 7). * P
    Control Gfp Expressing Adenovirus Adgfp, supplied by Vector Biolabs, used in various techniques. Bioz Stars score: 85/100, based on 23 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa gfp encoding mammalian expression vector
    S655 phosphorylation is a targeting signal for retrieval of <t>APP</t> to the Golgi . (A) Wt, S655A and S655E <t>APP-GFP</t> at the cell surface were labelled (0 min) by incubating COS-7 cells with the 22C11 anti-APP ectodomain antibody (
    Gfp Encoding Mammalian Expression Vector, supplied by TaKaRa, used in various techniques. Bioz Stars score: 85/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    InvivoGen gfp control vector
    S655 phosphorylation is a targeting signal for retrieval of <t>APP</t> to the Golgi . (A) Wt, S655A and S655E <t>APP-GFP</t> at the cell surface were labelled (0 min) by incubating COS-7 cells with the 22C11 anti-APP ectodomain antibody (
    Gfp Control Vector, supplied by InvivoGen, used in various techniques. Bioz Stars score: 88/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Lonza gfp control vector
    Mutation or SNO of C144 inhibits oxidant-induced cell death <t>HEK-293</t> cells expressing either <t>GFP-tagged</t> TRIM72 WT or TRIM72 C144S , or GFP alone were treated for (A) 3 hours or (B) 24 hours with H 2 O 2 (200 μmol/L) with or without GSNO pre-treatment (1 mmol/L, 1 hour). Cell viability was assayed by fluorescent imaging of ethidium bromide staining normalized to cell number (*p
    Gfp Control Vector, supplied by Lonza, used in various techniques. Bioz Stars score: 88/100, based on 33 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Lonza positive control gfp expression vector pmaxgfp
    Mutation or SNO of C144 inhibits oxidant-induced cell death <t>HEK-293</t> cells expressing either <t>GFP-tagged</t> TRIM72 WT or TRIM72 C144S , or GFP alone were treated for (A) 3 hours or (B) 24 hours with H 2 O 2 (200 μmol/L) with or without GSNO pre-treatment (1 mmol/L, 1 hour). Cell viability was assayed by fluorescent imaging of ethidium bromide staining normalized to cell number (*p
    Positive Control Gfp Expression Vector Pmaxgfp, supplied by Lonza, used in various techniques. Bioz Stars score: 90/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Novartis gfp control vector
    Mutation or SNO of C144 inhibits oxidant-induced cell death <t>HEK-293</t> cells expressing either <t>GFP-tagged</t> TRIM72 WT or TRIM72 C144S , or GFP alone were treated for (A) 3 hours or (B) 24 hours with H 2 O 2 (200 μmol/L) with or without GSNO pre-treatment (1 mmol/L, 1 hour). Cell viability was assayed by fluorescent imaging of ethidium bromide staining normalized to cell number (*p
    Gfp Control Vector, supplied by Novartis, used in various techniques. Bioz Stars score: 88/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Genechem control vector gfp
    Overexpression of <t>PNUTS</t> induces EMT and significantly increases the mobility and invasiveness of CNE-2 cells. Notes: ( A ) CNE-2 cells were infected with <t>ad-GFP</t> or ad-PNUTS at an efficiency measured by Western blotting. Western blotting was used to detect the expression of vimentin, N-cadherin, and E-cadherin. The internal control was GAPDH. ( B , C ) After overexpressing PNUTS, wound-healing assays (100× magnification) and Transwell assays (200× magnification) were used to detect the mobility and invasiveness of CNE-2 cells. Mean ± SD. n=3. * P
    Control Vector Gfp, supplied by Genechem, used in various techniques. Bioz Stars score: 93/100, based on 15 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    OriGene gfp expressing plasmid
    Overexpression of <t>PNUTS</t> induces EMT and significantly increases the mobility and invasiveness of CNE-2 cells. Notes: ( A ) CNE-2 cells were infected with <t>ad-GFP</t> or ad-PNUTS at an efficiency measured by Western blotting. Western blotting was used to detect the expression of vimentin, N-cadherin, and E-cadherin. The internal control was GAPDH. ( B , C ) After overexpressing PNUTS, wound-healing assays (100× magnification) and Transwell assays (200× magnification) were used to detect the mobility and invasiveness of CNE-2 cells. Mean ± SD. n=3. * P
    Gfp Expressing Plasmid, supplied by OriGene, used in various techniques. Bioz Stars score: 89/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Vector Biolabs adenovirus expressing gfp
    Overexpression of <t>PNUTS</t> induces EMT and significantly increases the mobility and invasiveness of CNE-2 cells. Notes: ( A ) CNE-2 cells were infected with <t>ad-GFP</t> or ad-PNUTS at an efficiency measured by Western blotting. Western blotting was used to detect the expression of vimentin, N-cadherin, and E-cadherin. The internal control was GAPDH. ( B , C ) After overexpressing PNUTS, wound-healing assays (100× magnification) and Transwell assays (200× magnification) were used to detect the mobility and invasiveness of CNE-2 cells. Mean ± SD. n=3. * P
    Adenovirus Expressing Gfp, supplied by Vector Biolabs, used in various techniques. Bioz Stars score: 88/100, based on 73 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Expression of GFP and Foxc2 genes after transfection. (A) Transfection efficiency at various MOIs (10, 50, 100 and 200) 72 hours after transfection (×100). (B) Cell proliferation after transfection in the Lv-Foxc2 and Lv-GFP groups detected using MTT assay. (C) Foxc2 gene expression measured by real-time PCR 72 hours after transfection. GAPDH was used as a housekeeping gene. (D) Foxc2 protein expression evaluated by western blot 72 hours after transfection. Results were expressed as mean±SD of triplicate experiments. #p

    Journal: BMC Musculoskeletal Disorders

    Article Title: Foxc2 regulates osteogenesis and angiogenesis of bone marrow mesenchymal stem cells

    doi: 10.1186/1471-2474-14-199

    Figure Lengend Snippet: Expression of GFP and Foxc2 genes after transfection. (A) Transfection efficiency at various MOIs (10, 50, 100 and 200) 72 hours after transfection (×100). (B) Cell proliferation after transfection in the Lv-Foxc2 and Lv-GFP groups detected using MTT assay. (C) Foxc2 gene expression measured by real-time PCR 72 hours after transfection. GAPDH was used as a housekeeping gene. (D) Foxc2 protein expression evaluated by western blot 72 hours after transfection. Results were expressed as mean±SD of triplicate experiments. #p

    Article Snippet: Lentiviral production and transfection A lentiviral vector expressing human Foxc2 and a control vector expressing GFP were purchased from Shanghai GeneChem Co., China.

    Techniques: Expressing, Transfection, MTT Assay, Real-time Polymerase Chain Reaction, Western Blot

    The effect of Foxc2 on osteogenic differentiation. (A , B and C) BMSCs were transfected with Lv-GFP or Lv-Foxc2 for 2 weeks. Cells stained with Alizarin red S ( A and B , a1 presents the Lv-Foxc2 group, and a2 presents the Lv-GFP group). Cells stained with ALP ( C , c1 presents the Lv-Foxc2 group and c2 presents the Lv-GFP group). (D) ALP activities in BMSCs. (E) Expressions of CON and Runx2 mRNA detected by real-time PCR 72 hours after transfection. The relative density of each gene was statistically analyzed. #p

    Journal: BMC Musculoskeletal Disorders

    Article Title: Foxc2 regulates osteogenesis and angiogenesis of bone marrow mesenchymal stem cells

    doi: 10.1186/1471-2474-14-199

    Figure Lengend Snippet: The effect of Foxc2 on osteogenic differentiation. (A , B and C) BMSCs were transfected with Lv-GFP or Lv-Foxc2 for 2 weeks. Cells stained with Alizarin red S ( A and B , a1 presents the Lv-Foxc2 group, and a2 presents the Lv-GFP group). Cells stained with ALP ( C , c1 presents the Lv-Foxc2 group and c2 presents the Lv-GFP group). (D) ALP activities in BMSCs. (E) Expressions of CON and Runx2 mRNA detected by real-time PCR 72 hours after transfection. The relative density of each gene was statistically analyzed. #p

    Article Snippet: Lentiviral production and transfection A lentiviral vector expressing human Foxc2 and a control vector expressing GFP were purchased from Shanghai GeneChem Co., China.

    Techniques: Transfection, Staining, ALP Assay, Real-time Polymerase Chain Reaction

    Foxc2 overexpression induces activation of ERK and PI3K in BMSCs. (A , B) BMSCs were transfected with Lv-GFP, Lv-Foxc2 or Lv-Foxc2 following pretreatment with 50 μM PD98059 or 10 μM LY294002 for 1 h. Cell lysates harvested after 2 weeks of culture were analyzed by SDS-PAGE and Western blot with anti ERK or anti-PI3K antibody. The result of ERK or PI3K expression was indicated by the ratio of band intensity of ERK or PI3K with β-actin. *p

    Journal: BMC Musculoskeletal Disorders

    Article Title: Foxc2 regulates osteogenesis and angiogenesis of bone marrow mesenchymal stem cells

    doi: 10.1186/1471-2474-14-199

    Figure Lengend Snippet: Foxc2 overexpression induces activation of ERK and PI3K in BMSCs. (A , B) BMSCs were transfected with Lv-GFP, Lv-Foxc2 or Lv-Foxc2 following pretreatment with 50 μM PD98059 or 10 μM LY294002 for 1 h. Cell lysates harvested after 2 weeks of culture were analyzed by SDS-PAGE and Western blot with anti ERK or anti-PI3K antibody. The result of ERK or PI3K expression was indicated by the ratio of band intensity of ERK or PI3K with β-actin. *p

    Article Snippet: Lentiviral production and transfection A lentiviral vector expressing human Foxc2 and a control vector expressing GFP were purchased from Shanghai GeneChem Co., China.

    Techniques: Over Expression, Activation Assay, Transfection, SDS Page, Western Blot, Expressing

    Western Blot of WT GFP and GFP Y39TAG mutant expression in a cell-free translation system. Synthesis of WT GFP and the GFP Y39TAG mutant was performed using the RTS E. coli HY Kit, to which the corresponding plasmid (500 µg/mL), purified Mj TyrRS and cognate suppressor Mj tRNA CUA (tRNA) or T-stem modified tRNA CUA Opt (denoted as *) were added. (A) Expression of WT GFP and the GFP Y39TAG mutant in the presence of Mj TyrRS (300 µg/mL) and synthetic Mj tRNA CUA (60 µg/mL). The band at 28 kDa corresponds to full-length GFP. (B) Western blot analysis demonstrates enhanced GFP Y39TAG protein expression as a function of increased Mj TyrRS concentrations in a cell-free reaction medium supplied with Mj tRNA CUA (60 µg/mL – top panel and 450 µg/mL – bottom panel). ( C ) Dependence of GFP Y39TAG yield on the type and concentration of nonsense suppressor, as visualized by Western blot.

    Journal: PLoS ONE

    Article Title: Enhanced Yield of Recombinant Proteins with Site-Specifically Incorporated Unnatural Amino Acids Using a Cell-Free Expression System

    doi: 10.1371/journal.pone.0068363

    Figure Lengend Snippet: Western Blot of WT GFP and GFP Y39TAG mutant expression in a cell-free translation system. Synthesis of WT GFP and the GFP Y39TAG mutant was performed using the RTS E. coli HY Kit, to which the corresponding plasmid (500 µg/mL), purified Mj TyrRS and cognate suppressor Mj tRNA CUA (tRNA) or T-stem modified tRNA CUA Opt (denoted as *) were added. (A) Expression of WT GFP and the GFP Y39TAG mutant in the presence of Mj TyrRS (300 µg/mL) and synthetic Mj tRNA CUA (60 µg/mL). The band at 28 kDa corresponds to full-length GFP. (B) Western blot analysis demonstrates enhanced GFP Y39TAG protein expression as a function of increased Mj TyrRS concentrations in a cell-free reaction medium supplied with Mj tRNA CUA (60 µg/mL – top panel and 450 µg/mL – bottom panel). ( C ) Dependence of GFP Y39TAG yield on the type and concentration of nonsense suppressor, as visualized by Western blot.

    Article Snippet: GFP Mutants The X-ray crystallographic structure of template GFP (PDB accession number: 1EMA), encoded by the GFP control vector (RTS, 5 PRIME, Hamburg, Germany), was analyzed to choose sites for UAA incorporation.

    Techniques: Western Blot, Mutagenesis, Expressing, Plasmid Preparation, Purification, Modification, Concentration Assay

    a A luciferase reporter expression assay was performed to examine whether miR-520d-5p actually targets TEAD1 and GATAD2B . Out of the more than ten predicted binding sites in the 3′UTRs of TEAD1 and GATAD2B , two sequences [TTACAAG and TACAAAG in the 3′UTR of TEAD1 ( top ) or GATAD2B ( bottom )] were chosen to test for miR-520d-5p binding. These sites were predicted based on the four databases described above. b A luciferase reporter expression assay revealed that miR-520d-5p bound to both the 3′UTR of TEAD1 and the 3′UTR of GATAD2B . In contrast, a mismatch of miR-520d-5p (a control for standardizing data) and a scramble group did not significantly bind to the 3′UTR sites, resulting in a failure to induce luciferase expression. Using a luciferase reporter expression assay, the potent sites of miR-520d-5p binding to the 3′UTR were identified and representatively shown at base pairs 4,569–4,581 and 7,463–7,776 in the 3′UTR of TEAD1 and 3,622–3,639 in the 3′UTR of GATAD2B , respectively. The minus signs (−) in the synthesized miR-520d-5p sequence represent the mismatches in the synthesized miR-520d-3p, and the minus signs in the control vector pMIRNA1/GFP sequence represent the mismatch sequences within miR-520d-5p that was expressed from pMIRNA1-520d-5p/GFP ( n = 4). The data were analyzed by t test (* P

    Journal: Drugs in R & D

    Article Title: Hsa-miR-520d Converts Fibroblasts into CD105+ Populations

    doi: 10.1007/s40268-014-0064-6

    Figure Lengend Snippet: a A luciferase reporter expression assay was performed to examine whether miR-520d-5p actually targets TEAD1 and GATAD2B . Out of the more than ten predicted binding sites in the 3′UTRs of TEAD1 and GATAD2B , two sequences [TTACAAG and TACAAAG in the 3′UTR of TEAD1 ( top ) or GATAD2B ( bottom )] were chosen to test for miR-520d-5p binding. These sites were predicted based on the four databases described above. b A luciferase reporter expression assay revealed that miR-520d-5p bound to both the 3′UTR of TEAD1 and the 3′UTR of GATAD2B . In contrast, a mismatch of miR-520d-5p (a control for standardizing data) and a scramble group did not significantly bind to the 3′UTR sites, resulting in a failure to induce luciferase expression. Using a luciferase reporter expression assay, the potent sites of miR-520d-5p binding to the 3′UTR were identified and representatively shown at base pairs 4,569–4,581 and 7,463–7,776 in the 3′UTR of TEAD1 and 3,622–3,639 in the 3′UTR of GATAD2B , respectively. The minus signs (−) in the synthesized miR-520d-5p sequence represent the mismatches in the synthesized miR-520d-3p, and the minus signs in the control vector pMIRNA1/GFP sequence represent the mismatch sequences within miR-520d-5p that was expressed from pMIRNA1-520d-5p/GFP ( n = 4). The data were analyzed by t test (* P

    Article Snippet: Synthesized miR-520d-5p (MBL, Nagoya, Japan) or the control vector pMIRNA1/GFP (System Biosciences, Mountain View, CA, USA) were co-transfected into NHDF cells together with each prepared luciferase expression vector (firefly or Renilla ).

    Techniques: Luciferase, Expressing, Binding Assay, Synthesized, Sequencing, Plasmid Preparation

    dCas9 3A3L or dCas9 3A targeting prevents senescence and increases proliferation. a , b Proliferation was assessed using a colorimetric assay 10, 15 and 20 days after targeting dCas9 3A3L to HIC1 , RASSF1 , PTEN and CDKN2A and magnetic sorting in primary myoepithelial cells from donors 1, 2 and 3. Dotted line represents the average absorbance from three dCas9 3A3LΔ targeted wells at each timepoint. The graphs show the average difference in absorbance as fold change compared to 3A3LΔ average (mean ± SEM, n = 3). c Cumulative population doublings over time for untransfected primary myoepithelial cells and those transfected with dCas9 3A3L or 3A3L∆ targeting the four genes without magnetic sorting from donor one. Fifty thousand cells were seeded at the start of each passage and cells counted after 5 days (mean ± SEM, n = 3, where error bars are smaller than points plotted they are not shown). ( d ) Light microscopy images of β-gal staining from untransfected (top left) primary myoepithelial cells from donor 1 and those transfected with dCas9 3A3L (bottom two) or 3A3L∆ (top right) targeting the four genes without magnetic sorting 20 days after transfection, blue/green colouring shows β-gal +ve cells; scale bar represents 125 µm. e Proliferation was assessed using a colorimetric assay 15, 20 and 25 days after targeting the four genes with dCas9 3A3L (red), dCas9 3A (orange), dCas9 3A-C706A-3L (yellow), dCas9 3A3L∆ (green) or dCas9m4 (pale green), or targeting only dCas9 3A3L without gRNAs (blue) without magnetic sorting using donor 1 myoepithelial cells. The data is shown as fold change compared to the average absorbance from 3 wells without cells (mean ± SEM, n = 3). f The percentage of GFP +ve cells after infection with no virus (green), hTERT with GFP marker (pink) or GFP control (purple). At infection day 0 cells are 38 days post-transfection with dCas9 3A3L targeting the four genes (mean ± SEM; n = 3, error bars are not shown where smaller than points plotted)

    Journal: Nature Communications

    Article Title: Hit-and-run epigenetic editing prevents senescence entry in primary breast cells from healthy donors

    doi: 10.1038/s41467-017-01078-2

    Figure Lengend Snippet: dCas9 3A3L or dCas9 3A targeting prevents senescence and increases proliferation. a , b Proliferation was assessed using a colorimetric assay 10, 15 and 20 days after targeting dCas9 3A3L to HIC1 , RASSF1 , PTEN and CDKN2A and magnetic sorting in primary myoepithelial cells from donors 1, 2 and 3. Dotted line represents the average absorbance from three dCas9 3A3LΔ targeted wells at each timepoint. The graphs show the average difference in absorbance as fold change compared to 3A3LΔ average (mean ± SEM, n = 3). c Cumulative population doublings over time for untransfected primary myoepithelial cells and those transfected with dCas9 3A3L or 3A3L∆ targeting the four genes without magnetic sorting from donor one. Fifty thousand cells were seeded at the start of each passage and cells counted after 5 days (mean ± SEM, n = 3, where error bars are smaller than points plotted they are not shown). ( d ) Light microscopy images of β-gal staining from untransfected (top left) primary myoepithelial cells from donor 1 and those transfected with dCas9 3A3L (bottom two) or 3A3L∆ (top right) targeting the four genes without magnetic sorting 20 days after transfection, blue/green colouring shows β-gal +ve cells; scale bar represents 125 µm. e Proliferation was assessed using a colorimetric assay 15, 20 and 25 days after targeting the four genes with dCas9 3A3L (red), dCas9 3A (orange), dCas9 3A-C706A-3L (yellow), dCas9 3A3L∆ (green) or dCas9m4 (pale green), or targeting only dCas9 3A3L without gRNAs (blue) without magnetic sorting using donor 1 myoepithelial cells. The data is shown as fold change compared to the average absorbance from 3 wells without cells (mean ± SEM, n = 3). f The percentage of GFP +ve cells after infection with no virus (green), hTERT with GFP marker (pink) or GFP control (purple). At infection day 0 cells are 38 days post-transfection with dCas9 3A3L targeting the four genes (mean ± SEM; n = 3, error bars are not shown where smaller than points plotted)

    Article Snippet: A retroviral vector expressing hTERT with a GFP marker (Addgene #69809) and a GFP-only control vector (Addgene #21654) were transfected into the Plat-A cells using jetprime.

    Techniques: Colorimetric Assay, Transfection, Light Microscopy, Staining, Infection, Marker

    VGF mRNA and eGFP expression patterns are similar following rAAV-VGF infusio n into the hypothalamus of both Siberian hamsters and mice . Representative pictures of eGFP expression (left) and VGF mRNA expression via in situ hybridisation (right) in the hypothalamus of Siberian hamsters (A and B) and mice (C and D) receiving bilateral injections of either rAAV-GFP (A and C) or rAAV-VGF (B and D) directed to the PVN. Scale bar = 2.5 mm.

    Journal: Journal of Neuroscience Methods

    Article Title: The use of a viral 2A sequence for the simultaneous over-expression of both the vgf gene and enhanced green fluorescent protein (eGFP) in vitro and in vivo

    doi: 10.1016/j.jneumeth.2015.08.013

    Figure Lengend Snippet: VGF mRNA and eGFP expression patterns are similar following rAAV-VGF infusio n into the hypothalamus of both Siberian hamsters and mice . Representative pictures of eGFP expression (left) and VGF mRNA expression via in situ hybridisation (right) in the hypothalamus of Siberian hamsters (A and B) and mice (C and D) receiving bilateral injections of either rAAV-GFP (A and C) or rAAV-VGF (B and D) directed to the PVN. Scale bar = 2.5 mm.

    Article Snippet: The rAAV-GFP control vector was purchased from Vector Biolabs (PA, USA); this had a viral titre of 1 × 1013 gc/ml and expressed eGFP under the control of a cytomegalovirus (CMV) promoter.

    Techniques: Expressing, Mouse Assay, In Situ, Hybridization

    A schematic to show steps leading to the production of rAAV-VGF . Full length mouse cDNA was isolated and inserted into the blunt cloning vector pSC-B-AMP/KAN for amplification using modified primers to produce the VGF-2A fragment. This product was cloned into the pAAV-CBA-AgRP-IRES-GFP backbone (previously used in Jethwa et al., 2010 ), following excision of the AgRP-IRES complex to produce pAAV-CBA-VGF-2a-GFP (pAAV-VGF-GFP). This was used for the in vitro checking (Study 1), before then being used to produce recombinant AAV-2 (rAAV) by Vector BioLabs (USA) yielding rAAV-VGF for in vivo infusion (study 2). ITR—internal tandem repeat, CMV/IE—cytomegalovirus intermediate early promoter, CBA—chicken β-actin promoter, eGFP—enhanced green fluorescent protein, WPRE—Woodchuck Hepatitis virus post-transcriptional regulatory element.

    Journal: Journal of Neuroscience Methods

    Article Title: The use of a viral 2A sequence for the simultaneous over-expression of both the vgf gene and enhanced green fluorescent protein (eGFP) in vitro and in vivo

    doi: 10.1016/j.jneumeth.2015.08.013

    Figure Lengend Snippet: A schematic to show steps leading to the production of rAAV-VGF . Full length mouse cDNA was isolated and inserted into the blunt cloning vector pSC-B-AMP/KAN for amplification using modified primers to produce the VGF-2A fragment. This product was cloned into the pAAV-CBA-AgRP-IRES-GFP backbone (previously used in Jethwa et al., 2010 ), following excision of the AgRP-IRES complex to produce pAAV-CBA-VGF-2a-GFP (pAAV-VGF-GFP). This was used for the in vitro checking (Study 1), before then being used to produce recombinant AAV-2 (rAAV) by Vector BioLabs (USA) yielding rAAV-VGF for in vivo infusion (study 2). ITR—internal tandem repeat, CMV/IE—cytomegalovirus intermediate early promoter, CBA—chicken β-actin promoter, eGFP—enhanced green fluorescent protein, WPRE—Woodchuck Hepatitis virus post-transcriptional regulatory element.

    Article Snippet: The rAAV-GFP control vector was purchased from Vector Biolabs (PA, USA); this had a viral titre of 1 × 1013 gc/ml and expressed eGFP under the control of a cytomegalovirus (CMV) promoter.

    Techniques: Isolation, Clone Assay, Plasmid Preparation, Amplification, Modification, Crocin Bleaching Assay, In Vitro, Recombinant, In Vivo

    Use of the viral 2A sequence in the rAAV-VGF construct results in widespread eGFP expression in the hypothalamus . Representative pictures of eGFP expression in the hypothalamus of Siberian hamsters and mice which received bilateral (200 nl) injections of either rAAV-GFP or rAAV-VGF directed to the PVN. (A–D) eGFP expression following infusion of rAAV-GFP control vector in the whole hypothalamus of (A) Siberian hamsters and (B) mice and in the cell soma and processes of the PVN of (C) Siberian hamsters and (D) mice. (E–G) eGFP expression following infusion of the rAAV-VGF vector in the hypothalamus of (E) Siberian hamsters and (F) mice and in the cell soma and processes of the PVN of (G) Siberian hamsters and (H) mice. Scale bar = 2.5 mm.

    Journal: Journal of Neuroscience Methods

    Article Title: The use of a viral 2A sequence for the simultaneous over-expression of both the vgf gene and enhanced green fluorescent protein (eGFP) in vitro and in vivo

    doi: 10.1016/j.jneumeth.2015.08.013

    Figure Lengend Snippet: Use of the viral 2A sequence in the rAAV-VGF construct results in widespread eGFP expression in the hypothalamus . Representative pictures of eGFP expression in the hypothalamus of Siberian hamsters and mice which received bilateral (200 nl) injections of either rAAV-GFP or rAAV-VGF directed to the PVN. (A–D) eGFP expression following infusion of rAAV-GFP control vector in the whole hypothalamus of (A) Siberian hamsters and (B) mice and in the cell soma and processes of the PVN of (C) Siberian hamsters and (D) mice. (E–G) eGFP expression following infusion of the rAAV-VGF vector in the hypothalamus of (E) Siberian hamsters and (F) mice and in the cell soma and processes of the PVN of (G) Siberian hamsters and (H) mice. Scale bar = 2.5 mm.

    Article Snippet: The rAAV-GFP control vector was purchased from Vector Biolabs (PA, USA); this had a viral titre of 1 × 1013 gc/ml and expressed eGFP under the control of a cytomegalovirus (CMV) promoter.

    Techniques: Sequencing, Construct, Expressing, Mouse Assay, Plasmid Preparation

    Decreased dendritic complexity in DSCAM-overexpressing hippocampal neurons. A , Representative micrographs of hippocampal neurons transfected at DIV7 with the indicated constructs. Neuronal morphology was visualized by GFP immunocytochemistry 2 d after transfection. Dendrites were identified as MAP2-positive neurites. Scale bars, 60 μm. B , Sholl analysis of the neurons transfected with GFP or DSCAM-IRES-GFP vectors. C , Total dendritic length of neurons transfected with the indicated constructs. D , Number of primary dendrites of neurons transfected with the indicated constructs. E , Somatic area of neurons transfected with the indicated plasmids. Mean values are shown (12–15 neurons); the error bars indicate the SEM, and asterisks denote statistically significant differences: * p

    Journal: The Journal of Neuroscience

    Article Title: NMDA-Mediated Regulation of DSCAM Dendritic Local Translation Is Lost in a Mouse Model of Down's Syndrome

    doi: 10.1523/JNEUROSCI.3457-10.2010

    Figure Lengend Snippet: Decreased dendritic complexity in DSCAM-overexpressing hippocampal neurons. A , Representative micrographs of hippocampal neurons transfected at DIV7 with the indicated constructs. Neuronal morphology was visualized by GFP immunocytochemistry 2 d after transfection. Dendrites were identified as MAP2-positive neurites. Scale bars, 60 μm. B , Sholl analysis of the neurons transfected with GFP or DSCAM-IRES-GFP vectors. C , Total dendritic length of neurons transfected with the indicated constructs. D , Number of primary dendrites of neurons transfected with the indicated constructs. E , Somatic area of neurons transfected with the indicated plasmids. Mean values are shown (12–15 neurons); the error bars indicate the SEM, and asterisks denote statistically significant differences: * p

    Article Snippet: A total of 0.8 μg of a plasmid expressing mouse DSCAM along with green fluorescent protein (GFP) protein (CMV-DSCAM-IRES-GFP) ( ) or a control vector expressing GFP protein (pEGFP-C1; Clontech) was transfected per well.

    Techniques: Transfection, Construct, Immunocytochemistry

    BATF silencing enhanced cytokine secretions by PPD-specific T cells in PBMCs in patients with ATB. (A) Representative fluorescence microscopy image of transfected primary human PBMCs after 24 h of electroporation with 2 μg GFP Vector, and red arrow indicated transfected PBMCs containing GFP (× 200). (B) Transfection efficiency of primary human PBMCs 24 h post electroporation were assessed by analysis of GFP protein expression by flow cytometry. (C) Silencing of BATF by a BATF siRNA SMART pool in primary human PBMCs from ATB patients measured by real-time quantitative PCR. Expression normalized to a housekeeping gene ( GAPDH ) is presented as fold change relative to negative control siRNA. (D) PBMCs were electroporated with indicated siRNA, then cultured with M. tb -specific antigen PPD (25 μg/mL) overnight, and IL-2 or IFN-γ mRNA levels were measured by real-time quantitative PCR. The information of controls and cell viability after transfection was provided in the Methods section BATF Small Interfering RNA (siRNA) Knockdown in ATB Patients ( 11 ). Data are expressed as mean ± SEM ( n = 7). * P

    Journal: Frontiers in Immunology

    Article Title: BATF Potentially Mediates Negative Regulation of PD-1/PD-Ls Pathway on T Cell Functions in Mycobacterium tuberculosis Infection

    doi: 10.3389/fimmu.2019.02430

    Figure Lengend Snippet: BATF silencing enhanced cytokine secretions by PPD-specific T cells in PBMCs in patients with ATB. (A) Representative fluorescence microscopy image of transfected primary human PBMCs after 24 h of electroporation with 2 μg GFP Vector, and red arrow indicated transfected PBMCs containing GFP (× 200). (B) Transfection efficiency of primary human PBMCs 24 h post electroporation were assessed by analysis of GFP protein expression by flow cytometry. (C) Silencing of BATF by a BATF siRNA SMART pool in primary human PBMCs from ATB patients measured by real-time quantitative PCR. Expression normalized to a housekeeping gene ( GAPDH ) is presented as fold change relative to negative control siRNA. (D) PBMCs were electroporated with indicated siRNA, then cultured with M. tb -specific antigen PPD (25 μg/mL) overnight, and IL-2 or IFN-γ mRNA levels were measured by real-time quantitative PCR. The information of controls and cell viability after transfection was provided in the Methods section BATF Small Interfering RNA (siRNA) Knockdown in ATB Patients ( 11 ). Data are expressed as mean ± SEM ( n = 7). * P

    Article Snippet: Per 5 × 106 PBMCs from ATB patients were resuspended in 100 μL room-temperature balanced Nucleofector®solution (Human T Cell Nucleofector®Kit, Lonza, Germany), and added with 200 nM siRNA (ON-TARGET plus Non-targeting pool or BATF ON-TARGET plus SMART pool, GE Dharmacon, USA) or 2 μg GFP Vector (Lonza, Germany).

    Techniques: Fluorescence, Microscopy, Transfection, Electroporation, Plasmid Preparation, Expressing, Flow Cytometry, Cytometry, Real-time Polymerase Chain Reaction, Negative Control, Cell Culture, Small Interfering RNA

    S655 phosphorylation is a targeting signal for retrieval of APP to the Golgi . (A) Wt, S655A and S655E APP-GFP at the cell surface were labelled (0 min) by incubating COS-7 cells with the 22C11 anti-APP ectodomain antibody (

    Journal: Molecular Neurodegeneration

    Article Title: Retrieval of the Alzheimer's amyloid precursor protein from the endosome to the TGN is S655 phosphorylation state-dependent and retromer-mediated

    doi: 10.1186/1750-1326-5-40

    Figure Lengend Snippet: S655 phosphorylation is a targeting signal for retrieval of APP to the Golgi . (A) Wt, S655A and S655E APP-GFP at the cell surface were labelled (0 min) by incubating COS-7 cells with the 22C11 anti-APP ectodomain antibody ("22C11 Uptake", Texas red staining) at 4°C. Following 15 and 30 min of incubation at 37°C, the localization of APP-GFP endocytic vesicles (green/red-22C11 co-localizing vesicles) were monitored. Arrows indicate the Golgi area and APP-GFP/22C11 endocytic vesicles co-localizing to or around the same region. ROI - region of interest denoting the Golgi area and co-localizing vesicles. (B) The highly green-fluorescent perinuclear structure in APP-GFP transfected cells was confirmed to be the Golgi in Wt APP-GFP and CFP-Golgi co-transfected cells, upon 30 min of 22C11 uptake. Bar, 10 μm. (C) confocal quantitative analysis of the degree of co-localization between uptaken 22C11 and the APP-GPP population at the Golgi area at 30 min of 22C11 antibody uptake (AU) at 37°C. Values are mean ± SEM, n = 20 cells. Statistical significance symbols used were: (*) for comparison of S655 phosphomutant and Wt data; ( + ) S655A vs S655E data. Statistical significance levels are presented as (**), for p

    Article Snippet: The resultant fragments were digested with endonucleases (Age I and Nru I) and subcloned into the Age I/Sma I restriction sites of the GFP-encoding mammalian expression vector (pEGFP-N1, Clontech) as N-terminal APP-GFP translational fusions.

    Techniques: Staining, Incubation, Transfection

    VPS35 down-regulation impairs S655E APP-GFP retrieval to the TGN and decreases its turnover rate . (A) Left: Immunodetection of VPS35 levels in COS-7 cells co-transfected with N1 EGFP and several concentrations of anti-VPS35 siRNAs for 24 h. Tubulin was probed as a control. Right: Quantification of VPS35 levels, plotted as percentages of control conditions (cells only transfected with N1 EGFP). Values are mean ± SEM (n = 2-6); (#), statistical significance of p ≤ 0.05 for siRNA VPS35 vs control data. (B) The 15 min 22C11 antibody uptake assay (Alexa350 blue staining) was applied to monitor S655E APP-GFP protein retrieval from the endosome to the TGN upon COS-7 cells pre-incubation with 5nM VPS35 siRNAs for 24 h to downregulate VPS35 (Texas red staining). (B.I) Control: cells only transfected with S655E APP-GFP. (B.II - B.III) siRNA VPS35: cells co-transfected with S655E APP-GFP and VPS35 siRNAs; in the majority of the population, the APP-GFP signal at the Golgi was not visible (B.II ), while in a minor percentage of cells APP-GFP was still visible at the Golgi (B.III) . Bar, 10 μm. (C) S655E APP-GFP turnover rate in CHX upon cells pre-incubation with 5nM VPS35 siRNAs for 24 h. Left: Immunoblot analysis of COS-7 transfected cells lysates using the anti-APP 22C11 and anti-tubulin antibodies. Bands a and b, immature and mature APP-GFP forms, respectively, migrating above endogenous APP forms. Right: mature S655E levels were quantified and plotted as percentages of OD values at 0 h in CHX. Values are mean ± SEM (n = 3); (#), statistical significance of p ≤ 0.05 for siRNA VPS35 plus vs control data.

    Journal: Molecular Neurodegeneration

    Article Title: Retrieval of the Alzheimer's amyloid precursor protein from the endosome to the TGN is S655 phosphorylation state-dependent and retromer-mediated

    doi: 10.1186/1750-1326-5-40

    Figure Lengend Snippet: VPS35 down-regulation impairs S655E APP-GFP retrieval to the TGN and decreases its turnover rate . (A) Left: Immunodetection of VPS35 levels in COS-7 cells co-transfected with N1 EGFP and several concentrations of anti-VPS35 siRNAs for 24 h. Tubulin was probed as a control. Right: Quantification of VPS35 levels, plotted as percentages of control conditions (cells only transfected with N1 EGFP). Values are mean ± SEM (n = 2-6); (#), statistical significance of p ≤ 0.05 for siRNA VPS35 vs control data. (B) The 15 min 22C11 antibody uptake assay (Alexa350 blue staining) was applied to monitor S655E APP-GFP protein retrieval from the endosome to the TGN upon COS-7 cells pre-incubation with 5nM VPS35 siRNAs for 24 h to downregulate VPS35 (Texas red staining). (B.I) Control: cells only transfected with S655E APP-GFP. (B.II - B.III) siRNA VPS35: cells co-transfected with S655E APP-GFP and VPS35 siRNAs; in the majority of the population, the APP-GFP signal at the Golgi was not visible (B.II ), while in a minor percentage of cells APP-GFP was still visible at the Golgi (B.III) . Bar, 10 μm. (C) S655E APP-GFP turnover rate in CHX upon cells pre-incubation with 5nM VPS35 siRNAs for 24 h. Left: Immunoblot analysis of COS-7 transfected cells lysates using the anti-APP 22C11 and anti-tubulin antibodies. Bands a and b, immature and mature APP-GFP forms, respectively, migrating above endogenous APP forms. Right: mature S655E levels were quantified and plotted as percentages of OD values at 0 h in CHX. Values are mean ± SEM (n = 3); (#), statistical significance of p ≤ 0.05 for siRNA VPS35 plus vs control data.

    Article Snippet: The resultant fragments were digested with endonucleases (Age I and Nru I) and subcloned into the Age I/Sma I restriction sites of the GFP-encoding mammalian expression vector (pEGFP-N1, Clontech) as N-terminal APP-GFP translational fusions.

    Techniques: Immunodetection, Transfection, Staining, Incubation

    Co-immunoprecipitation of APP, VPS35 and SorLA in COS-7 and HEK293 cells . (A) Endogenous APP and VPS35 were co-immunoprecipitated in COS-7 cells using the anti-APP N-terminus 22C11 and the anti-VPS35 antibodies (αVPS35). Negative controls (C) were performed by immunoprecipitating cells with the same secondary antibodies, sepharose- (IP VPS35 control) and agarose- (IP 22C11) linked, respectively. (B) Transfected Wt APP-GFP and endogenous VPS35 co-immunoprecipitate from COS-7 cells using the indicated antibodies (Ab). (C) Transfected Wt, S655A and S655E APP-GFP were immunoprecipitated in COS-7 cells with the anti-GFP antibody, and the co-immunoprecipitated endogenous VPS35 forms were detected with an anti-VPS35 antibody. Asterisk (*), non-specific IgGs bands (IgGs raised in goat) of the IP samples. Non-transfected cells (C(nt)) submitted to the same IP procedures (incubated with primary and agarose-linked secondary antibodies) were used as control in (B) and (C) . (D) HEK293T cells were cotransfected with SorLA cDNA, APP 695 and eGFP (transfection control). Immunoprecipitation was performed using antibodies raised against the C-terminus of APP (369), the N-terminus of SorLA (αSorLA) or using pre-immune serum as negative controls (C). Immunoprecipitation and co-immunoprecipitation were detected by western blot (WB) using the anti-APP 369 antibody and an anti-SorLA C-terminus antibody (αSorLA C-term). Immunoblot analysis included GFP as an additional negative control.

    Journal: Molecular Neurodegeneration

    Article Title: Retrieval of the Alzheimer's amyloid precursor protein from the endosome to the TGN is S655 phosphorylation state-dependent and retromer-mediated

    doi: 10.1186/1750-1326-5-40

    Figure Lengend Snippet: Co-immunoprecipitation of APP, VPS35 and SorLA in COS-7 and HEK293 cells . (A) Endogenous APP and VPS35 were co-immunoprecipitated in COS-7 cells using the anti-APP N-terminus 22C11 and the anti-VPS35 antibodies (αVPS35). Negative controls (C) were performed by immunoprecipitating cells with the same secondary antibodies, sepharose- (IP VPS35 control) and agarose- (IP 22C11) linked, respectively. (B) Transfected Wt APP-GFP and endogenous VPS35 co-immunoprecipitate from COS-7 cells using the indicated antibodies (Ab). (C) Transfected Wt, S655A and S655E APP-GFP were immunoprecipitated in COS-7 cells with the anti-GFP antibody, and the co-immunoprecipitated endogenous VPS35 forms were detected with an anti-VPS35 antibody. Asterisk (*), non-specific IgGs bands (IgGs raised in goat) of the IP samples. Non-transfected cells (C(nt)) submitted to the same IP procedures (incubated with primary and agarose-linked secondary antibodies) were used as control in (B) and (C) . (D) HEK293T cells were cotransfected with SorLA cDNA, APP 695 and eGFP (transfection control). Immunoprecipitation was performed using antibodies raised against the C-terminus of APP (369), the N-terminus of SorLA (αSorLA) or using pre-immune serum as negative controls (C). Immunoprecipitation and co-immunoprecipitation were detected by western blot (WB) using the anti-APP 369 antibody and an anti-SorLA C-terminus antibody (αSorLA C-term). Immunoblot analysis included GFP as an additional negative control.

    Article Snippet: The resultant fragments were digested with endonucleases (Age I and Nru I) and subcloned into the Age I/Sma I restriction sites of the GFP-encoding mammalian expression vector (pEGFP-N1, Clontech) as N-terminal APP-GFP translational fusions.

    Techniques: Immunoprecipitation, Transfection, Incubation, Western Blot, Negative Control

    A pool of endocytic APP-GFP is targeted to the TGN . Wt APP-GFP at COS-7 cell surface was labelled by incubating cells with the 22C11 anti-APP ectodomain antibody (

    Journal: Molecular Neurodegeneration

    Article Title: Retrieval of the Alzheimer's amyloid precursor protein from the endosome to the TGN is S655 phosphorylation state-dependent and retromer-mediated

    doi: 10.1186/1750-1326-5-40

    Figure Lengend Snippet: A pool of endocytic APP-GFP is targeted to the TGN . Wt APP-GFP at COS-7 cell surface was labelled by incubating cells with the 22C11 anti-APP ectodomain antibody ("22C11 Uptake", Texas red staining) at 4°C. At 0 min, 22C11 staining is maintained at the cell surface (microphotographs taken at the plasma membrane focal plane). Following 15 min at 37°C, part of the 22C11 population can be observed at vesicular structures around the Golgi (arrowhead; microphotographs taken at the Golgi focal plane). Endocytic 22C11/APP-GFP targeting to the TGN was confirmed by co-localization with a pECFP-Golgi construct ("CFP-Golgi", blue fluorescence), a construct targeted to the trans and medial region of the Golgi. Of note, since the CFP-Golgi fusion protein has a weak blue fluorescence, it is virtually non-visible at the plasma membrane focal plane. ROI - region of interest, 2.0-fold magnified. Several white vesicles, denoting 22C11/APP-GFP/CFP-Golgi co-localization, can be observed at the TGN (arrows in ROIs), A tubular morphology can be depicted for some of these vesicles in the ROIs. Bar, 10 μm.

    Article Snippet: The resultant fragments were digested with endonucleases (Age I and Nru I) and subcloned into the Age I/Sma I restriction sites of the GFP-encoding mammalian expression vector (pEGFP-N1, Clontech) as N-terminal APP-GFP translational fusions.

    Techniques: Staining, Construct, Fluorescence

    Endocytosed APP is present in Clathrin and Rab 5-positive vesicles tubulating vesicles . The 22C11 uptake assay (Alexa350 blue staining) was repeated in COS-7 cells for further characterization of the vesicles retrieving endocytosed APP-GFP to the TGN. Immunocytochemistry analysis using antibodies against (A) clathrin and (B) Rab 5 (Texas red staining) confirmed the presence of these proteins in APP-GFP endocytic vesicles. ROI, region of interest, 1.7- and 2.0-fold magnified, respectively. Arrows in ROIs depict APP tubulating endosomes, further magnified. While 22C11/APP-GFP and clathrin were observed to be sorted from the vacuole to the emerging tubule, Rab 5 appears to be maintained in the vacuole. Bar, 10 μm. (C) confocal quantitative analysis of the degree of co-localization between Wt APP-GPP and uptaken 22C11 with clathrin and Rab-5. Values are mean ± SEM, n = 20 cells.

    Journal: Molecular Neurodegeneration

    Article Title: Retrieval of the Alzheimer's amyloid precursor protein from the endosome to the TGN is S655 phosphorylation state-dependent and retromer-mediated

    doi: 10.1186/1750-1326-5-40

    Figure Lengend Snippet: Endocytosed APP is present in Clathrin and Rab 5-positive vesicles tubulating vesicles . The 22C11 uptake assay (Alexa350 blue staining) was repeated in COS-7 cells for further characterization of the vesicles retrieving endocytosed APP-GFP to the TGN. Immunocytochemistry analysis using antibodies against (A) clathrin and (B) Rab 5 (Texas red staining) confirmed the presence of these proteins in APP-GFP endocytic vesicles. ROI, region of interest, 1.7- and 2.0-fold magnified, respectively. Arrows in ROIs depict APP tubulating endosomes, further magnified. While 22C11/APP-GFP and clathrin were observed to be sorted from the vacuole to the emerging tubule, Rab 5 appears to be maintained in the vacuole. Bar, 10 μm. (C) confocal quantitative analysis of the degree of co-localization between Wt APP-GPP and uptaken 22C11 with clathrin and Rab-5. Values are mean ± SEM, n = 20 cells.

    Article Snippet: The resultant fragments were digested with endonucleases (Age I and Nru I) and subcloned into the Age I/Sma I restriction sites of the GFP-encoding mammalian expression vector (pEGFP-N1, Clontech) as N-terminal APP-GFP translational fusions.

    Techniques: Staining, Immunocytochemistry

    S655 APP phosphomutants exhibit differential cellular catabolism . (A) Wt and S655 phosphomutants turnover rates in COS-7 cells. Upper panel: Immunoblot analysis of APP-GFP transfected cells lysates using the anti-APP antibody 22C11. Bands a and b, immature (N-glycosylated) and mature (N- and O-glycosylated) APP-GFP forms, respectively, migrating above endogenous APP forms of ~115 kD and ~109 kD (potentially immature APP 751/770 and APP 695 , respectively). APP-GFP bands identity was further confirmed using an anti-GFP antibody (data not shown). Lower panel: plotted data of the levels of immature (left graph) and mature (right graph) APP-GFP, expressed as percentage of OD values at 0 h in CHX. Values are mean ± SEM (n = 6). Statistical significance symbols: (*), S655 phosphomutants vs. Wt; ( + ), S655A vs. S655E; statistical significance levels are presented as (*/ + ) for p ≤ 0.05; ( ++ ), for p

    Journal: Molecular Neurodegeneration

    Article Title: Retrieval of the Alzheimer's amyloid precursor protein from the endosome to the TGN is S655 phosphorylation state-dependent and retromer-mediated

    doi: 10.1186/1750-1326-5-40

    Figure Lengend Snippet: S655 APP phosphomutants exhibit differential cellular catabolism . (A) Wt and S655 phosphomutants turnover rates in COS-7 cells. Upper panel: Immunoblot analysis of APP-GFP transfected cells lysates using the anti-APP antibody 22C11. Bands a and b, immature (N-glycosylated) and mature (N- and O-glycosylated) APP-GFP forms, respectively, migrating above endogenous APP forms of ~115 kD and ~109 kD (potentially immature APP 751/770 and APP 695 , respectively). APP-GFP bands identity was further confirmed using an anti-GFP antibody (data not shown). Lower panel: plotted data of the levels of immature (left graph) and mature (right graph) APP-GFP, expressed as percentage of OD values at 0 h in CHX. Values are mean ± SEM (n = 6). Statistical significance symbols: (*), S655 phosphomutants vs. Wt; ( + ), S655A vs. S655E; statistical significance levels are presented as (*/ + ) for p ≤ 0.05; ( ++ ), for p

    Article Snippet: The resultant fragments were digested with endonucleases (Age I and Nru I) and subcloned into the Age I/Sma I restriction sites of the GFP-encoding mammalian expression vector (pEGFP-N1, Clontech) as N-terminal APP-GFP translational fusions.

    Techniques: Transfection

    S655 phosphorylation-dependent APP retrieval occurs via VPS35-containing endocytic vesicles . (A) APP-GFP transfected COS-7 cells were subjected to the 15 min 22C11 uptake assay (Alexa350 blue staining) and the co-localization of endocytosed APP-GFP proteins (blue/green vesicles) with the VPS35 retromer subunit (Texas red staining) was monitored. The Golgi area denotes the highest degree of co-localization, and a region was magnified for better visualization of the tubulating vesicles (ROI-4.0-fold magnified; arrows point to APP-GFP/AU22C11/VPS35-positive structures for Wt and S655E, and arrowhead to a S655A APP-GFP-negative, AU22C11/VPS35-positive structure). Some of the tubulating vesicles for Wt and S655E have been represented schematically with a black line. (B) Co-localization between APP-GFP or 22C11 uptake with VPS35 was quantified using confocal software. Values are mean ± SEM, n = 30 cells. Statistical significance symbols: (*), S655 phosphomutant vs Wt data; ( + ), S655A vs S655E data. Statistically significant levels are presented as (**) for p

    Journal: Molecular Neurodegeneration

    Article Title: Retrieval of the Alzheimer's amyloid precursor protein from the endosome to the TGN is S655 phosphorylation state-dependent and retromer-mediated

    doi: 10.1186/1750-1326-5-40

    Figure Lengend Snippet: S655 phosphorylation-dependent APP retrieval occurs via VPS35-containing endocytic vesicles . (A) APP-GFP transfected COS-7 cells were subjected to the 15 min 22C11 uptake assay (Alexa350 blue staining) and the co-localization of endocytosed APP-GFP proteins (blue/green vesicles) with the VPS35 retromer subunit (Texas red staining) was monitored. The Golgi area denotes the highest degree of co-localization, and a region was magnified for better visualization of the tubulating vesicles (ROI-4.0-fold magnified; arrows point to APP-GFP/AU22C11/VPS35-positive structures for Wt and S655E, and arrowhead to a S655A APP-GFP-negative, AU22C11/VPS35-positive structure). Some of the tubulating vesicles for Wt and S655E have been represented schematically with a black line. (B) Co-localization between APP-GFP or 22C11 uptake with VPS35 was quantified using confocal software. Values are mean ± SEM, n = 30 cells. Statistical significance symbols: (*), S655 phosphomutant vs Wt data; ( + ), S655A vs S655E data. Statistically significant levels are presented as (**) for p

    Article Snippet: The resultant fragments were digested with endonucleases (Age I and Nru I) and subcloned into the Age I/Sma I restriction sites of the GFP-encoding mammalian expression vector (pEGFP-N1, Clontech) as N-terminal APP-GFP translational fusions.

    Techniques: Transfection, Staining, Software

    APP-GFP traverses the endocytic pathway . (a) COS-7 cells transfected with Wt APP-GFP were exposed for 3 h to the protein synthesis inhibitor CHX and incubated in the last 15 min with Texas Red-conjugated transferring. Co-localization (yellow/orange fluorescence vesicles) between transferrin (Texas red endocytic vesicles) and APP-GFP green cytoplasmic vesicles can be observed (Overlay panel). Immunocytochemistry analysis with (b) an early (Rab 5) and (c) late (Rab 7) endosomal markers were also performed and co-localization observed. ROI, region of interest, denotes APP-GFP co-localizing endocytic vesicles. Bar, 10 μm. Fluorescence intensity profiles are also presented, representing the voxels through the white arrowed lines indicated in the ROI overlay images; asterisks denote co-localizing vesicles. A microphotograph of the eGFP protein is presented as a negative control, GFP alone is distributed through the nucleus and cytoplasm, but is not targeted to cytoplasmic vesicles.

    Journal: Molecular Neurodegeneration

    Article Title: Retrieval of the Alzheimer's amyloid precursor protein from the endosome to the TGN is S655 phosphorylation state-dependent and retromer-mediated

    doi: 10.1186/1750-1326-5-40

    Figure Lengend Snippet: APP-GFP traverses the endocytic pathway . (a) COS-7 cells transfected with Wt APP-GFP were exposed for 3 h to the protein synthesis inhibitor CHX and incubated in the last 15 min with Texas Red-conjugated transferring. Co-localization (yellow/orange fluorescence vesicles) between transferrin (Texas red endocytic vesicles) and APP-GFP green cytoplasmic vesicles can be observed (Overlay panel). Immunocytochemistry analysis with (b) an early (Rab 5) and (c) late (Rab 7) endosomal markers were also performed and co-localization observed. ROI, region of interest, denotes APP-GFP co-localizing endocytic vesicles. Bar, 10 μm. Fluorescence intensity profiles are also presented, representing the voxels through the white arrowed lines indicated in the ROI overlay images; asterisks denote co-localizing vesicles. A microphotograph of the eGFP protein is presented as a negative control, GFP alone is distributed through the nucleus and cytoplasm, but is not targeted to cytoplasmic vesicles.

    Article Snippet: The resultant fragments were digested with endonucleases (Age I and Nru I) and subcloned into the Age I/Sma I restriction sites of the GFP-encoding mammalian expression vector (pEGFP-N1, Clontech) as N-terminal APP-GFP translational fusions.

    Techniques: Transfection, Incubation, Transferring, Fluorescence, Immunocytochemistry, Negative Control

    Mutation or SNO of C144 inhibits oxidant-induced cell death HEK-293 cells expressing either GFP-tagged TRIM72 WT or TRIM72 C144S , or GFP alone were treated for (A) 3 hours or (B) 24 hours with H 2 O 2 (200 μmol/L) with or without GSNO pre-treatment (1 mmol/L, 1 hour). Cell viability was assayed by fluorescent imaging of ethidium bromide staining normalized to cell number (*p

    Journal: Journal of molecular and cellular cardiology

    Article Title: S-nitrosylation of TRIM72 at cysteine 144 is critical for protection against oxidation-induced protein degradation and cell death

    doi: 10.1016/j.yjmcc.2014.01.010

    Figure Lengend Snippet: Mutation or SNO of C144 inhibits oxidant-induced cell death HEK-293 cells expressing either GFP-tagged TRIM72 WT or TRIM72 C144S , or GFP alone were treated for (A) 3 hours or (B) 24 hours with H 2 O 2 (200 μmol/L) with or without GSNO pre-treatment (1 mmol/L, 1 hour). Cell viability was assayed by fluorescent imaging of ethidium bromide staining normalized to cell number (*p

    Article Snippet: HEK-293 cells were transfected with GFP-tagged TRIM72WT , TRIM72C144S or a GFP control vector as described above.

    Techniques: Mutagenesis, Expressing, Imaging, Staining

    Enhanced miR-125b expression and reduced CCL4 expression in naïve CD8 T cells. (A) Enhanced miR-125b level (∽78-fold) in naïve CD8 T cells transfected with GFP-tagged miR-125b expressing vector compared to naïve CD8 T cells transfected with GFP control vector. Twelve hours post-transfection, GFP-positive cells were isolated by cell sorting for determining the levels of miR-125b. (B) Reduced expression of CCL4 mRNA ( N = 9) and (C) protein ( N = 12) in miR-125b enriched naïve CD8 T cells as compared to control cells. Supernatants were collected after 16 h stimulation with anti-CD3 and anti-CD28 from transfected cells. The data were log transformed and presented as mean and SEM. Paired Student t-test was used (** P

    Journal: Aging Cell

    Article Title: MicroRNA-125b modulates inflammatory chemokine CCL4 expression in immune cells and its reduction causes CCL4 increase with age

    doi: 10.1111/acel.12294

    Figure Lengend Snippet: Enhanced miR-125b expression and reduced CCL4 expression in naïve CD8 T cells. (A) Enhanced miR-125b level (∽78-fold) in naïve CD8 T cells transfected with GFP-tagged miR-125b expressing vector compared to naïve CD8 T cells transfected with GFP control vector. Twelve hours post-transfection, GFP-positive cells were isolated by cell sorting for determining the levels of miR-125b. (B) Reduced expression of CCL4 mRNA ( N = 9) and (C) protein ( N = 12) in miR-125b enriched naïve CD8 T cells as compared to control cells. Supernatants were collected after 16 h stimulation with anti-CD3 and anti-CD28 from transfected cells. The data were log transformed and presented as mean and SEM. Paired Student t-test was used (** P

    Article Snippet: 0.4 μg/million cell of GFP plus miR-125b vector or control GFP vector was transfected into freshly isolated naïve CD8 T cells using Human T cell Nucleofector kit (Lonza, Allendale, NJ, USA).

    Techniques: Expressing, Transfection, Plasmid Preparation, Isolation, FACS, Transformation Assay

    Overexpression of PNUTS induces EMT and significantly increases the mobility and invasiveness of CNE-2 cells. Notes: ( A ) CNE-2 cells were infected with ad-GFP or ad-PNUTS at an efficiency measured by Western blotting. Western blotting was used to detect the expression of vimentin, N-cadherin, and E-cadherin. The internal control was GAPDH. ( B , C ) After overexpressing PNUTS, wound-healing assays (100× magnification) and Transwell assays (200× magnification) were used to detect the mobility and invasiveness of CNE-2 cells. Mean ± SD. n=3. * P

    Journal: OncoTargets and therapy

    Article Title: PNUTS mediates ionizing radiation-induced CNE-2 nasopharyngeal carcinoma cell migration, invasion, and epithelial–mesenchymal transition via the PI3K/AKT signaling pathway

    doi: 10.2147/OTT.S188571

    Figure Lengend Snippet: Overexpression of PNUTS induces EMT and significantly increases the mobility and invasiveness of CNE-2 cells. Notes: ( A ) CNE-2 cells were infected with ad-GFP or ad-PNUTS at an efficiency measured by Western blotting. Western blotting was used to detect the expression of vimentin, N-cadherin, and E-cadherin. The internal control was GAPDH. ( B , C ) After overexpressing PNUTS, wound-healing assays (100× magnification) and Transwell assays (200× magnification) were used to detect the mobility and invasiveness of CNE-2 cells. Mean ± SD. n=3. * P

    Article Snippet: Adenovirus infection An adenovirus expressing PNUTS and its control vector GFP were packaged and synthesized by Genechem (Shanghai, China).

    Techniques: Over Expression, Infection, Western Blot, Expressing