retroviral transduction Search Results


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  • 99
    Thermo Fisher retroviral transduction 293ft cells
    Retroviral Transduction 293ft Cells, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Addgene inc retroviral transduction
    Vector for <t>retroviral</t> transduction of BCRs. Diagram of the retroviral vector for ectopic expression of different isotypes of BCR in primary B cells and B cell lines. LTR, long terminal repeat, which on the 3’ end is self-inactivating (white x); ψ + , extended packaging signal; EF1, EF1α promoter, Amp R , ampicillin resistantance gene. DOI: http://dx.doi.org/10.7554/eLife.21238.004
    Retroviral Transduction, supplied by Addgene inc, used in various techniques. Bioz Stars score: 92/100, based on 246 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa retroviral transduction
    Gli1 + cells in bone marrow fibrosis are transcriptionally distinct from Gli1 + cells in homeostasis. Bigenic Gli1CreER;tdTomato mice were injected with tamoxifen (3x10mg p.o.) at 8 weeks of age, lethally irradiated at 10 days after the last tamoxifen dose and received c-kit enriched hematopoietic stem cells from wildtype littermates expressing either Thrombopoietin-cDNA (ThPO, n=5, 3 males) or control-cDNA (control, n=4, 3 males; both lentiviral SFFV-iGFP vector backbone). Mice were sacrificed at 70 days after transplantation. Gli1 + cells were sort-purified as lin - GFP - dtTomato + and subjected to RNA sequencing. (A) Principal component analysis (PCA). (B) Heatmap representation with hierarchical clustering (C) Significantly directional enriched (median FDR > 0.05) pathways. (D) Fragments per kilobase of exon per million fragments mapped (FPKM) values of CXCL4 in control and ThPO group; mean±SEM. (E) Cytokine analysis in serum and cytokine-free medium supernatant of ckit+ cells expressing ThPO or control (lentiviral SFFV-iGFP vector backbone) or Jak2(V617F) or control (both MCSV-IRES-GFP <t>retroviral</t> backbone). n=2. (F) FPKM value of stromal derived factor 1 (SDF 1/ CXCL12) in control and ThPO group; mean±SEM. (G) Representative images of bone marrow from Gli1CreER t2 ;tdTomato mice transplanted with bone marrow from wildtype littermates expressing either ThPO or control-cDNA stained with the neutral lipid staining LipidTOX. Scale bars 50μm. Arrow indicating LipidTOX positive Gli1 + cells. (H) .
    Retroviral Transduction, supplied by TaKaRa, used in various techniques. Bioz Stars score: 92/100, based on 704 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Addgene inc pmxs retroviral transduction
    Gli1 + cells in bone marrow fibrosis are transcriptionally distinct from Gli1 + cells in homeostasis. Bigenic Gli1CreER;tdTomato mice were injected with tamoxifen (3x10mg p.o.) at 8 weeks of age, lethally irradiated at 10 days after the last tamoxifen dose and received c-kit enriched hematopoietic stem cells from wildtype littermates expressing either Thrombopoietin-cDNA (ThPO, n=5, 3 males) or control-cDNA (control, n=4, 3 males; both lentiviral SFFV-iGFP vector backbone). Mice were sacrificed at 70 days after transplantation. Gli1 + cells were sort-purified as lin - GFP - dtTomato + and subjected to RNA sequencing. (A) Principal component analysis (PCA). (B) Heatmap representation with hierarchical clustering (C) Significantly directional enriched (median FDR > 0.05) pathways. (D) Fragments per kilobase of exon per million fragments mapped (FPKM) values of CXCL4 in control and ThPO group; mean±SEM. (E) Cytokine analysis in serum and cytokine-free medium supernatant of ckit+ cells expressing ThPO or control (lentiviral SFFV-iGFP vector backbone) or Jak2(V617F) or control (both MCSV-IRES-GFP <t>retroviral</t> backbone). n=2. (F) FPKM value of stromal derived factor 1 (SDF 1/ CXCL12) in control and ThPO group; mean±SEM. (G) Representative images of bone marrow from Gli1CreER t2 ;tdTomato mice transplanted with bone marrow from wildtype littermates expressing either ThPO or control-cDNA stained with the neutral lipid staining LipidTOX. Scale bars 50μm. Arrow indicating LipidTOX positive Gli1 + cells. (H) .
    Pmxs Retroviral Transduction, supplied by Addgene inc, used in various techniques. Bioz Stars score: 85/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    DSMZ retroviral transduction retroviral transduction
    Gli1 + cells in bone marrow fibrosis are transcriptionally distinct from Gli1 + cells in homeostasis. Bigenic Gli1CreER;tdTomato mice were injected with tamoxifen (3x10mg p.o.) at 8 weeks of age, lethally irradiated at 10 days after the last tamoxifen dose and received c-kit enriched hematopoietic stem cells from wildtype littermates expressing either Thrombopoietin-cDNA (ThPO, n=5, 3 males) or control-cDNA (control, n=4, 3 males; both lentiviral SFFV-iGFP vector backbone). Mice were sacrificed at 70 days after transplantation. Gli1 + cells were sort-purified as lin - GFP - dtTomato + and subjected to RNA sequencing. (A) Principal component analysis (PCA). (B) Heatmap representation with hierarchical clustering (C) Significantly directional enriched (median FDR > 0.05) pathways. (D) Fragments per kilobase of exon per million fragments mapped (FPKM) values of CXCL4 in control and ThPO group; mean±SEM. (E) Cytokine analysis in serum and cytokine-free medium supernatant of ckit+ cells expressing ThPO or control (lentiviral SFFV-iGFP vector backbone) or Jak2(V617F) or control (both MCSV-IRES-GFP <t>retroviral</t> backbone). n=2. (F) FPKM value of stromal derived factor 1 (SDF 1/ CXCL12) in control and ThPO group; mean±SEM. (G) Representative images of bone marrow from Gli1CreER t2 ;tdTomato mice transplanted with bone marrow from wildtype littermates expressing either ThPO or control-cDNA stained with the neutral lipid staining LipidTOX. Scale bars 50μm. Arrow indicating LipidTOX positive Gli1 + cells. (H) .
    Retroviral Transduction Retroviral Transduction, supplied by DSMZ, used in various techniques. Bioz Stars score: 90/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Becton Dickinson retroviral transduction
    Expression and localization of oncogenic NRAS and its PTM mutants . (A) Schematic diagram of <t>retroviral</t> expression vectors used to transduce NRASD12, NRASD12 C181S , or NRASD12 C186S . (B) Immunoblot of lysates of NIH3T3 cells stably expressing the vector control, NRASD12, NRASD12 C181S or NRASD12 C186S with a pan anti-RAS antibody (top band represents double Myc-tagged-NRAS; and bottom band, endogenous Ras). (C) NIH3T3 cells expressing GFP-fused NRASD12, NRASD12 C181S , or NRASD12 C186S , costained with fluorescence-conjugated antibodies against Golgi (Golga-7) or ER (BIP) resident proteins, were visualized on a Leica TCS SP2 Spectral Confocal Microscope (original magnification, ×630).
    Retroviral Transduction, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 93/100, based on 338 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Orbigen phoenixtm retroviral transduction system
    Expression and localization of oncogenic NRAS and its PTM mutants . (A) Schematic diagram of <t>retroviral</t> expression vectors used to transduce NRASD12, NRASD12 C181S , or NRASD12 C186S . (B) Immunoblot of lysates of NIH3T3 cells stably expressing the vector control, NRASD12, NRASD12 C181S or NRASD12 C186S with a pan anti-RAS antibody (top band represents double Myc-tagged-NRAS; and bottom band, endogenous Ras). (C) NIH3T3 cells expressing GFP-fused NRASD12, NRASD12 C181S , or NRASD12 C186S , costained with fluorescence-conjugated antibodies against Golgi (Golga-7) or ER (BIP) resident proteins, were visualized on a Leica TCS SP2 Spectral Confocal Microscope (original magnification, ×630).
    Phoenixtm Retroviral Transduction System, supplied by Orbigen, used in various techniques. Bioz Stars score: 88/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Greiner Bio retroviral transduction
    Expression and localization of oncogenic NRAS and its PTM mutants . (A) Schematic diagram of <t>retroviral</t> expression vectors used to transduce NRASD12, NRASD12 C181S , or NRASD12 C186S . (B) Immunoblot of lysates of NIH3T3 cells stably expressing the vector control, NRASD12, NRASD12 C181S or NRASD12 C186S with a pan anti-RAS antibody (top band represents double Myc-tagged-NRAS; and bottom band, endogenous Ras). (C) NIH3T3 cells expressing GFP-fused NRASD12, NRASD12 C181S , or NRASD12 C186S , costained with fluorescence-conjugated antibodies against Golgi (Golga-7) or ER (BIP) resident proteins, were visualized on a Leica TCS SP2 Spectral Confocal Microscope (original magnification, ×630).
    Retroviral Transduction, supplied by Greiner Bio, used in various techniques. Bioz Stars score: 93/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    STEMCELL Technologies Inc retroviral transduction
    Expression and localization of oncogenic NRAS and its PTM mutants . (A) Schematic diagram of <t>retroviral</t> expression vectors used to transduce NRASD12, NRASD12 C181S , or NRASD12 C186S . (B) Immunoblot of lysates of NIH3T3 cells stably expressing the vector control, NRASD12, NRASD12 C181S or NRASD12 C186S with a pan anti-RAS antibody (top band represents double Myc-tagged-NRAS; and bottom band, endogenous Ras). (C) NIH3T3 cells expressing GFP-fused NRASD12, NRASD12 C181S , or NRASD12 C186S , costained with fluorescence-conjugated antibodies against Golgi (Golga-7) or ER (BIP) resident proteins, were visualized on a Leica TCS SP2 Spectral Confocal Microscope (original magnification, ×630).
    Retroviral Transduction, supplied by STEMCELL Technologies Inc, used in various techniques. Bioz Stars score: 92/100, based on 102 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Addgene inc retroviral transduction vector pbabepuro
    <t>Retroviral</t> transduction of D2SC/1 cells . (A) D2SC/1 cells were cultured in the supernatant of the retroviral packaging cell line FLYA4lacZ3 for 24 h and LacZ transduced cells were visualized by X-gal staining. (B) The mp53 retroviral transduction vectors are illustrated. Predicted K[d] restricted peptide presentation is marked by a box. Amino acids at position 2, 3, 5, and carboxyl termini function as anchors or auxiliary anchors within the MHC pocket. (C) Flow cytometry of mp53-transduced D2SC/1. Stable amphotropic packaging cell lines were generated by calcium phosphate transfection of FLY-AF-13 cells with <t>pBABEpuro–p53C132F/E168G</t> or pBABEpuro–p53M234I and puromycin selection before D2SC/1 cells were transduced three times with 1.5–3 × 10 4 CFU/ml of budding virus for 15 h. Cells were fixed and stained with 1 μg/ml PAb 421 anti-p53 antibody. Filled histograms represent stained cells transduced with the pBABEpuro control vector, open histograms D2SC/1-p53C132F/E168G, or D2SC/1-p53M234I transduced cells.
    Retroviral Transduction Vector Pbabepuro, supplied by Addgene inc, used in various techniques. Bioz Stars score: 86/100, based on 15 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa standard retroviral transduction
    <t>Retroviral</t> transduction of D2SC/1 cells . (A) D2SC/1 cells were cultured in the supernatant of the retroviral packaging cell line FLYA4lacZ3 for 24 h and LacZ transduced cells were visualized by X-gal staining. (B) The mp53 retroviral transduction vectors are illustrated. Predicted K[d] restricted peptide presentation is marked by a box. Amino acids at position 2, 3, 5, and carboxyl termini function as anchors or auxiliary anchors within the MHC pocket. (C) Flow cytometry of mp53-transduced D2SC/1. Stable amphotropic packaging cell lines were generated by calcium phosphate transfection of FLY-AF-13 cells with <t>pBABEpuro–p53C132F/E168G</t> or pBABEpuro–p53M234I and puromycin selection before D2SC/1 cells were transduced three times with 1.5–3 × 10 4 CFU/ml of budding virus for 15 h. Cells were fixed and stained with 1 μg/ml PAb 421 anti-p53 antibody. Filled histograms represent stained cells transduced with the pBABEpuro control vector, open histograms D2SC/1-p53C132F/E168G, or D2SC/1-p53M234I transduced cells.
    Standard Retroviral Transduction, supplied by TaKaRa, used in various techniques. Bioz Stars score: 93/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Addgene inc retrovirus transduction
    <t>Retroviral</t> transduction of D2SC/1 cells . (A) D2SC/1 cells were cultured in the supernatant of the retroviral packaging cell line FLYA4lacZ3 for 24 h and LacZ transduced cells were visualized by X-gal staining. (B) The mp53 retroviral transduction vectors are illustrated. Predicted K[d] restricted peptide presentation is marked by a box. Amino acids at position 2, 3, 5, and carboxyl termini function as anchors or auxiliary anchors within the MHC pocket. (C) Flow cytometry of mp53-transduced D2SC/1. Stable amphotropic packaging cell lines were generated by calcium phosphate transfection of FLY-AF-13 cells with <t>pBABEpuro–p53C132F/E168G</t> or pBABEpuro–p53M234I and puromycin selection before D2SC/1 cells were transduced three times with 1.5–3 × 10 4 CFU/ml of budding virus for 15 h. Cells were fixed and stained with 1 μg/ml PAb 421 anti-p53 antibody. Filled histograms represent stained cells transduced with the pBABEpuro control vector, open histograms D2SC/1-p53C132F/E168G, or D2SC/1-p53M234I transduced cells.
    Retrovirus Transduction, supplied by Addgene inc, used in various techniques. Bioz Stars score: 89/100, based on 25 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Addgene inc retroviral transduction retroviral vectors plpc n myc
    <t>Retroviral</t> transduction of D2SC/1 cells . (A) D2SC/1 cells were cultured in the supernatant of the retroviral packaging cell line FLYA4lacZ3 for 24 h and LacZ transduced cells were visualized by X-gal staining. (B) The mp53 retroviral transduction vectors are illustrated. Predicted K[d] restricted peptide presentation is marked by a box. Amino acids at position 2, 3, 5, and carboxyl termini function as anchors or auxiliary anchors within the MHC pocket. (C) Flow cytometry of mp53-transduced D2SC/1. Stable amphotropic packaging cell lines were generated by calcium phosphate transfection of FLY-AF-13 cells with <t>pBABEpuro–p53C132F/E168G</t> or pBABEpuro–p53M234I and puromycin selection before D2SC/1 cells were transduced three times with 1.5–3 × 10 4 CFU/ml of budding virus for 15 h. Cells were fixed and stained with 1 μg/ml PAb 421 anti-p53 antibody. Filled histograms represent stained cells transduced with the pBABEpuro control vector, open histograms D2SC/1-p53C132F/E168G, or D2SC/1-p53M234I transduced cells.
    Retroviral Transduction Retroviral Vectors Plpc N Myc, supplied by Addgene inc, used in various techniques. Bioz Stars score: 88/100, based on 72 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Geneservice retroviral transduction mgc image
    <t>Retroviral</t> transduction of D2SC/1 cells . (A) D2SC/1 cells were cultured in the supernatant of the retroviral packaging cell line FLYA4lacZ3 for 24 h and LacZ transduced cells were visualized by X-gal staining. (B) The mp53 retroviral transduction vectors are illustrated. Predicted K[d] restricted peptide presentation is marked by a box. Amino acids at position 2, 3, 5, and carboxyl termini function as anchors or auxiliary anchors within the MHC pocket. (C) Flow cytometry of mp53-transduced D2SC/1. Stable amphotropic packaging cell lines were generated by calcium phosphate transfection of FLY-AF-13 cells with <t>pBABEpuro–p53C132F/E168G</t> or pBABEpuro–p53M234I and puromycin selection before D2SC/1 cells were transduced three times with 1.5–3 × 10 4 CFU/ml of budding virus for 15 h. Cells were fixed and stained with 1 μg/ml PAb 421 anti-p53 antibody. Filled histograms represent stained cells transduced with the pBABEpuro control vector, open histograms D2SC/1-p53C132F/E168G, or D2SC/1-p53M234I transduced cells.
    Retroviral Transduction Mgc Image, supplied by Geneservice, 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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    Addgene inc retrovirus transduction plncx akt vectors
    <t>AKT</t> is essential for the modulation of keratinocyte differentiation by HPV18 (A) Schematic showing the KGFR signalling pathway and the effects of the dominant active (DA) and negative (DN) AKT. HPV18 genome containing keratinocytes were infected with empty <t>retrovirus,</t> or with retroviruses encoding DA AKT (B) or DN AKT (C) . Cells were differentiated in high calcium media for 48 hours prior to lysis and analysed for expression of cytokeratin 1 (K1) to assess differentiation. Expression of the exogenous AKT was confirmed using an antibody against the HA epitope and GAPDH served as a loading control.
    Retrovirus Transduction Plncx Akt Vectors, supplied by Addgene inc, used in various techniques. Bioz Stars score: 91/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa retroviral vector transduction
    <t>AKT</t> is essential for the modulation of keratinocyte differentiation by HPV18 (A) Schematic showing the KGFR signalling pathway and the effects of the dominant active (DA) and negative (DN) AKT. HPV18 genome containing keratinocytes were infected with empty <t>retrovirus,</t> or with retroviruses encoding DA AKT (B) or DN AKT (C) . Cells were differentiated in high calcium media for 48 hours prior to lysis and analysed for expression of cytokeratin 1 (K1) to assess differentiation. Expression of the exogenous AKT was confirmed using an antibody against the HA epitope and GAPDH served as a loading control.
    Retroviral Vector Transduction, supplied by TaKaRa, used in various techniques. Bioz Stars score: 85/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Novus Biologicals retromax retroviral transduction assay
    <t>AKT</t> is essential for the modulation of keratinocyte differentiation by HPV18 (A) Schematic showing the KGFR signalling pathway and the effects of the dominant active (DA) and negative (DN) AKT. HPV18 genome containing keratinocytes were infected with empty <t>retrovirus,</t> or with retroviruses encoding DA AKT (B) or DN AKT (C) . Cells were differentiated in high calcium media for 48 hours prior to lysis and analysed for expression of cytokeratin 1 (K1) to assess differentiation. Expression of the exogenous AKT was confirmed using an antibody against the HA epitope and GAPDH served as a loading control.
    Retromax Retroviral Transduction Assay, supplied by Novus Biologicals, used in various techniques. Bioz Stars score: 85/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa retroviral transduction human
    NLRC3 regulates upstream signalling molecules within the PI3K–AKT–mTOR pathway a , Immunofluorescence staining of primary fibroblasts and frequency of co-localisation between mTOR and LAMP1 (n= > 150). b , Images and quantification of colon tumours in littermate Apc Min/+ and Apc Min/+ Nlrc3 −/− mice 40 days after treatment with vehicle or NVP-BEZ235. c , Immunohistochemical staining of colon tissues from mice treated as in b. d , Immunoprecipitation of the GFP tag in 239T cells transfected with a plasmid encoding GFP alone or NLRC3-GFP. e , Immunoblot of mouse colon tissues and densitometric quantification. f , Immunoblot of primary mouse fibroblasts transduced with a <t>retroviral</t> vector encoding GFP or NLRC3-GFP, with or without stimulation with IGF-1, and densitometric quantification. Scale bar, 10 µm ( a ), 200 µm ( c ). * P
    Retroviral Transduction Human, supplied by TaKaRa, used in various techniques. Bioz Stars score: 90/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Becton Dickinson retroviral transduction pe cy7 anti cd11b
    NLRC3 regulates upstream signalling molecules within the PI3K–AKT–mTOR pathway a , Immunofluorescence staining of primary fibroblasts and frequency of co-localisation between mTOR and LAMP1 (n= > 150). b , Images and quantification of colon tumours in littermate Apc Min/+ and Apc Min/+ Nlrc3 −/− mice 40 days after treatment with vehicle or NVP-BEZ235. c , Immunohistochemical staining of colon tissues from mice treated as in b. d , Immunoprecipitation of the GFP tag in 239T cells transfected with a plasmid encoding GFP alone or NLRC3-GFP. e , Immunoblot of mouse colon tissues and densitometric quantification. f , Immunoblot of primary mouse fibroblasts transduced with a <t>retroviral</t> vector encoding GFP or NLRC3-GFP, with or without stimulation with IGF-1, and densitometric quantification. Scale bar, 10 µm ( a ), 200 µm ( c ). * P
    Retroviral Transduction Pe Cy7 Anti Cd11b, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 85/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Addgene inc retrovirus transduction plncx akt vector
    NLRC3 regulates upstream signalling molecules within the PI3K–AKT–mTOR pathway a , Immunofluorescence staining of primary fibroblasts and frequency of co-localisation between mTOR and LAMP1 (n= > 150). b , Images and quantification of colon tumours in littermate Apc Min/+ and Apc Min/+ Nlrc3 −/− mice 40 days after treatment with vehicle or NVP-BEZ235. c , Immunohistochemical staining of colon tissues from mice treated as in b. d , Immunoprecipitation of the GFP tag in 239T cells transfected with a plasmid encoding GFP alone or NLRC3-GFP. e , Immunoblot of mouse colon tissues and densitometric quantification. f , Immunoblot of primary mouse fibroblasts transduced with a <t>retroviral</t> vector encoding GFP or NLRC3-GFP, with or without stimulation with IGF-1, and densitometric quantification. Scale bar, 10 µm ( a ), 200 µm ( c ). * P
    Retrovirus Transduction Plncx Akt Vector, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher retroviral transduction lipofectamine 2000
    NLRC3 regulates upstream signalling molecules within the PI3K–AKT–mTOR pathway a , Immunofluorescence staining of primary fibroblasts and frequency of co-localisation between mTOR and LAMP1 (n= > 150). b , Images and quantification of colon tumours in littermate Apc Min/+ and Apc Min/+ Nlrc3 −/− mice 40 days after treatment with vehicle or NVP-BEZ235. c , Immunohistochemical staining of colon tissues from mice treated as in b. d , Immunoprecipitation of the GFP tag in 239T cells transfected with a plasmid encoding GFP alone or NLRC3-GFP. e , Immunoblot of mouse colon tissues and densitometric quantification. f , Immunoblot of primary mouse fibroblasts transduced with a <t>retroviral</t> vector encoding GFP or NLRC3-GFP, with or without stimulation with IGF-1, and densitometric quantification. Scale bar, 10 µm ( a ), 200 µm ( c ). * P
    Retroviral Transduction Lipofectamine 2000, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Addgene inc retroviral transduction over expression
    Autophagic flux in virus-specific CD8 T cells is inversely correlated with cell proliferation status. ( a ) MSCV <t>retroviral</t> constructs showing the transgenes (GFP-LC3b and GFP-LC3b-G120A) introduced in P14 cells to probe for autophagy activity. ( b ) Flow cytometry plots of adoptively transferred P14 cells in spleens. P14 cells transduced with retroviral plasmid of either MIT-GFP-LC3b or MIT-G120A are positive for the congenic marker Thy1.1. Day 5, 8 and 30 p.i. splenocytes were used for the analysis. The percentage of GFP-negative cells out of the transduced P14 (Thy1.1 + ) cells from each group is highlighted in blue on the lower right corner of each plot and is summarized in ( c ). ( d ) Representative ImageStream ® data showing images of P14 cells from either GFP-LC3b or G120A groups on day 8 p.i.. Images showing GFP signals of GFP + MIT vector-transduced P14 cells (Ly5.1 + Thy1.1 + ). Summary graph of ImageStream ® analysis is plotted in ( e ) showing percentage of GFP + or Thy1.1 + P14 cells that exhibited more than one GFP punctum. Errors bars represent SEM. ( b ) and ( d ) are representative of three independent experiments, n≥3 mice in each group.
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    Vector for retroviral transduction of BCRs. Diagram of the retroviral vector for ectopic expression of different isotypes of BCR in primary B cells and B cell lines. LTR, long terminal repeat, which on the 3’ end is self-inactivating (white x); ψ + , extended packaging signal; EF1, EF1α promoter, Amp R , ampicillin resistantance gene. DOI: http://dx.doi.org/10.7554/eLife.21238.004

    Journal: eLife

    Article Title: Regulation of B cell fate by chronic activity of the IgE B cell receptor

    doi: 10.7554/eLife.21238

    Figure Lengend Snippet: Vector for retroviral transduction of BCRs. Diagram of the retroviral vector for ectopic expression of different isotypes of BCR in primary B cells and B cell lines. LTR, long terminal repeat, which on the 3’ end is self-inactivating (white x); ψ + , extended packaging signal; EF1, EF1α promoter, Amp R , ampicillin resistantance gene. DOI: http://dx.doi.org/10.7554/eLife.21238.004

    Article Snippet: To allow assessment of the expression of BCR constructs by retroviral transduction, the coding sequences of the fluorescent protein Cerulean ( ) (obtained from Addgene), light chain, and heavy chains of different isotypes of BCR were linked in-frame by T2A peptide sequences without internal stop codons and cloned downstream of the EF1α promoter using the restriction sites indicated in .

    Techniques: Plasmid Preparation, Transduction, Expressing

    Gli1 + cells in bone marrow fibrosis are transcriptionally distinct from Gli1 + cells in homeostasis. Bigenic Gli1CreER;tdTomato mice were injected with tamoxifen (3x10mg p.o.) at 8 weeks of age, lethally irradiated at 10 days after the last tamoxifen dose and received c-kit enriched hematopoietic stem cells from wildtype littermates expressing either Thrombopoietin-cDNA (ThPO, n=5, 3 males) or control-cDNA (control, n=4, 3 males; both lentiviral SFFV-iGFP vector backbone). Mice were sacrificed at 70 days after transplantation. Gli1 + cells were sort-purified as lin - GFP - dtTomato + and subjected to RNA sequencing. (A) Principal component analysis (PCA). (B) Heatmap representation with hierarchical clustering (C) Significantly directional enriched (median FDR > 0.05) pathways. (D) Fragments per kilobase of exon per million fragments mapped (FPKM) values of CXCL4 in control and ThPO group; mean±SEM. (E) Cytokine analysis in serum and cytokine-free medium supernatant of ckit+ cells expressing ThPO or control (lentiviral SFFV-iGFP vector backbone) or Jak2(V617F) or control (both MCSV-IRES-GFP retroviral backbone). n=2. (F) FPKM value of stromal derived factor 1 (SDF 1/ CXCL12) in control and ThPO group; mean±SEM. (G) Representative images of bone marrow from Gli1CreER t2 ;tdTomato mice transplanted with bone marrow from wildtype littermates expressing either ThPO or control-cDNA stained with the neutral lipid staining LipidTOX. Scale bars 50μm. Arrow indicating LipidTOX positive Gli1 + cells. (H) .

    Journal: Cell stem cell

    Article Title: Gli1+ mesenchymal stromal cells are a key driver of bone marrow fibrosis and an important cellular therapeutic target

    doi: 10.1016/j.stem.2017.03.008

    Figure Lengend Snippet: Gli1 + cells in bone marrow fibrosis are transcriptionally distinct from Gli1 + cells in homeostasis. Bigenic Gli1CreER;tdTomato mice were injected with tamoxifen (3x10mg p.o.) at 8 weeks of age, lethally irradiated at 10 days after the last tamoxifen dose and received c-kit enriched hematopoietic stem cells from wildtype littermates expressing either Thrombopoietin-cDNA (ThPO, n=5, 3 males) or control-cDNA (control, n=4, 3 males; both lentiviral SFFV-iGFP vector backbone). Mice were sacrificed at 70 days after transplantation. Gli1 + cells were sort-purified as lin - GFP - dtTomato + and subjected to RNA sequencing. (A) Principal component analysis (PCA). (B) Heatmap representation with hierarchical clustering (C) Significantly directional enriched (median FDR > 0.05) pathways. (D) Fragments per kilobase of exon per million fragments mapped (FPKM) values of CXCL4 in control and ThPO group; mean±SEM. (E) Cytokine analysis in serum and cytokine-free medium supernatant of ckit+ cells expressing ThPO or control (lentiviral SFFV-iGFP vector backbone) or Jak2(V617F) or control (both MCSV-IRES-GFP retroviral backbone). n=2. (F) FPKM value of stromal derived factor 1 (SDF 1/ CXCL12) in control and ThPO group; mean±SEM. (G) Representative images of bone marrow from Gli1CreER t2 ;tdTomato mice transplanted with bone marrow from wildtype littermates expressing either ThPO or control-cDNA stained with the neutral lipid staining LipidTOX. Scale bars 50μm. Arrow indicating LipidTOX positive Gli1 + cells. (H) .

    Article Snippet: Retroviral transduction was performed on retroNectin (Takara Bio) coated cell culture dishes loaded with unconcentrated virus.

    Techniques: Mouse Assay, Injection, Irradiation, Expressing, Plasmid Preparation, Transplantation Assay, Purification, RNA Sequencing Assay, Derivative Assay, Staining

    Gli1 + cells are myofibroblast precursors in JAK2(V617F) induced myelofibrosis and can be targeted pharmacologically by GANT61 (A) Bigenic Gli1CreER t2 ;tdTomato mice received tamoxifen (3x10mg p.o.) and were lethally irradiated 10 days after the last tamoxifen dose and transplanted with 2x10 5 c-kit purified bone marrow (BM) cells from wild type littermates that had been transduced with either control (n=10) or Jak2(V617F) (n=10) cDNA (both MCSV-IRES-GFP retroviral backbone vector). Mice were injected with GANT61 (50mg/kg body weight) or vehicle (corn oil / ethanol) every other day between 8 and 17 weeks after transplantation (control + vehicle n=3, 2 males; control+GANT61 n=5, 3 males; Jak2(V617F) + vehicle n=4, 3 males; Jak2(V617F) + GANT61 n=4, 3 males). (B) Grading of reticulin fibrosis. ***p

    Journal: Cell stem cell

    Article Title: Gli1+ mesenchymal stromal cells are a key driver of bone marrow fibrosis and an important cellular therapeutic target

    doi: 10.1016/j.stem.2017.03.008

    Figure Lengend Snippet: Gli1 + cells are myofibroblast precursors in JAK2(V617F) induced myelofibrosis and can be targeted pharmacologically by GANT61 (A) Bigenic Gli1CreER t2 ;tdTomato mice received tamoxifen (3x10mg p.o.) and were lethally irradiated 10 days after the last tamoxifen dose and transplanted with 2x10 5 c-kit purified bone marrow (BM) cells from wild type littermates that had been transduced with either control (n=10) or Jak2(V617F) (n=10) cDNA (both MCSV-IRES-GFP retroviral backbone vector). Mice were injected with GANT61 (50mg/kg body weight) or vehicle (corn oil / ethanol) every other day between 8 and 17 weeks after transplantation (control + vehicle n=3, 2 males; control+GANT61 n=5, 3 males; Jak2(V617F) + vehicle n=4, 3 males; Jak2(V617F) + GANT61 n=4, 3 males). (B) Grading of reticulin fibrosis. ***p

    Article Snippet: Retroviral transduction was performed on retroNectin (Takara Bio) coated cell culture dishes loaded with unconcentrated virus.

    Techniques: Mouse Assay, Irradiation, Purification, Transduction, Plasmid Preparation, Injection, Transplantation Assay

    The overexpression of EpCAM in HepG2 cells. (A) The overexpression of EpCAM was confirmed by Western blot analysis using anti-EPCAM antibody. (B) The cell-surface overexpression of EpCAM was confirmed by FACS analysis using anti-EpCAM antibody. The retroviral expression system was used to ectopically express EpCAM. The FLAG tag was inserted at the C-terminal end of EpCAM.

    Journal: Molecules and Cells

    Article Title: Identification of DNA Aptamers toward Epithelial Cell Adhesion Molecule via Cell-SELEX

    doi: 10.14348/molcells.2014.0208

    Figure Lengend Snippet: The overexpression of EpCAM in HepG2 cells. (A) The overexpression of EpCAM was confirmed by Western blot analysis using anti-EPCAM antibody. (B) The cell-surface overexpression of EpCAM was confirmed by FACS analysis using anti-EpCAM antibody. The retroviral expression system was used to ectopically express EpCAM. The FLAG tag was inserted at the C-terminal end of EpCAM.

    Article Snippet: EpCAM was overexpressed by using a retroviral transduction system (pRetroX-IRES DsRedvector, Clontech).

    Techniques: Over Expression, Western Blot, FACS, Expressing, FLAG-tag

    Expression and localization of oncogenic NRAS and its PTM mutants . (A) Schematic diagram of retroviral expression vectors used to transduce NRASD12, NRASD12 C181S , or NRASD12 C186S . (B) Immunoblot of lysates of NIH3T3 cells stably expressing the vector control, NRASD12, NRASD12 C181S or NRASD12 C186S with a pan anti-RAS antibody (top band represents double Myc-tagged-NRAS; and bottom band, endogenous Ras). (C) NIH3T3 cells expressing GFP-fused NRASD12, NRASD12 C181S , or NRASD12 C186S , costained with fluorescence-conjugated antibodies against Golgi (Golga-7) or ER (BIP) resident proteins, were visualized on a Leica TCS SP2 Spectral Confocal Microscope (original magnification, ×630).

    Journal: Blood

    Article Title: Palmitoylation of oncogenic NRAS is essential for leukemogenesis

    doi: 10.1182/blood-2009-03-213876

    Figure Lengend Snippet: Expression and localization of oncogenic NRAS and its PTM mutants . (A) Schematic diagram of retroviral expression vectors used to transduce NRASD12, NRASD12 C181S , or NRASD12 C186S . (B) Immunoblot of lysates of NIH3T3 cells stably expressing the vector control, NRASD12, NRASD12 C181S or NRASD12 C186S with a pan anti-RAS antibody (top band represents double Myc-tagged-NRAS; and bottom band, endogenous Ras). (C) NIH3T3 cells expressing GFP-fused NRASD12, NRASD12 C181S , or NRASD12 C186S , costained with fluorescence-conjugated antibodies against Golgi (Golga-7) or ER (BIP) resident proteins, were visualized on a Leica TCS SP2 Spectral Confocal Microscope (original magnification, ×630).

    Article Snippet: NIH3T3 and 32D cl-3 cell lines stably expressing NRASD12, NRASD12C181S , NRASD12C186S , or the control GFP were created by retroviral transduction as described., All cell lines were sorted by GFP expression to more than 95% homogeneity by fluorescence-activated cell sorting (FACS) using a FACSAria (BD Biosciences).

    Techniques: Expressing, Transduction, Stable Transfection, Plasmid Preparation, Fluorescence, Microscopy

    Retroviral transduction of D2SC/1 cells . (A) D2SC/1 cells were cultured in the supernatant of the retroviral packaging cell line FLYA4lacZ3 for 24 h and LacZ transduced cells were visualized by X-gal staining. (B) The mp53 retroviral transduction vectors are illustrated. Predicted K[d] restricted peptide presentation is marked by a box. Amino acids at position 2, 3, 5, and carboxyl termini function as anchors or auxiliary anchors within the MHC pocket. (C) Flow cytometry of mp53-transduced D2SC/1. Stable amphotropic packaging cell lines were generated by calcium phosphate transfection of FLY-AF-13 cells with pBABEpuro–p53C132F/E168G or pBABEpuro–p53M234I and puromycin selection before D2SC/1 cells were transduced three times with 1.5–3 × 10 4 CFU/ml of budding virus for 15 h. Cells were fixed and stained with 1 μg/ml PAb 421 anti-p53 antibody. Filled histograms represent stained cells transduced with the pBABEpuro control vector, open histograms D2SC/1-p53C132F/E168G, or D2SC/1-p53M234I transduced cells.

    Journal: Frontiers in Oncology

    Article Title: Adaptive Resistance to Immunotherapy Directed Against p53 Can be Overcome by Global Expression of Tumor-Antigens in Dendritic Cells

    doi: 10.3389/fonc.2014.00270

    Figure Lengend Snippet: Retroviral transduction of D2SC/1 cells . (A) D2SC/1 cells were cultured in the supernatant of the retroviral packaging cell line FLYA4lacZ3 for 24 h and LacZ transduced cells were visualized by X-gal staining. (B) The mp53 retroviral transduction vectors are illustrated. Predicted K[d] restricted peptide presentation is marked by a box. Amino acids at position 2, 3, 5, and carboxyl termini function as anchors or auxiliary anchors within the MHC pocket. (C) Flow cytometry of mp53-transduced D2SC/1. Stable amphotropic packaging cell lines were generated by calcium phosphate transfection of FLY-AF-13 cells with pBABEpuro–p53C132F/E168G or pBABEpuro–p53M234I and puromycin selection before D2SC/1 cells were transduced three times with 1.5–3 × 10 4 CFU/ml of budding virus for 15 h. Cells were fixed and stained with 1 μg/ml PAb 421 anti-p53 antibody. Filled histograms represent stained cells transduced with the pBABEpuro control vector, open histograms D2SC/1-p53C132F/E168G, or D2SC/1-p53M234I transduced cells.

    Article Snippet: The missense point mutations C132F, E168G, and M234I present in the p53 alleles were verified by sequence analysis. p53M234I and p53C132F/E168G were cloned into the retroviral transduction vector pBABEpuro (Addgene, Cambridge, MA, USA).

    Techniques: Transduction, Cell Culture, Staining, Flow Cytometry, Cytometry, Generated, Transfection, Selection, Plasmid Preparation

    AKT is essential for the modulation of keratinocyte differentiation by HPV18 (A) Schematic showing the KGFR signalling pathway and the effects of the dominant active (DA) and negative (DN) AKT. HPV18 genome containing keratinocytes were infected with empty retrovirus, or with retroviruses encoding DA AKT (B) or DN AKT (C) . Cells were differentiated in high calcium media for 48 hours prior to lysis and analysed for expression of cytokeratin 1 (K1) to assess differentiation. Expression of the exogenous AKT was confirmed using an antibody against the HA epitope and GAPDH served as a loading control.

    Journal: Oncotarget

    Article Title: Human papillomavirus type 18 E5 oncogene supports cell cycle progression and impairs epithelial differentiation by modulating growth factor receptor signalling during the virus life cycle

    doi: 10.18632/oncotarget.21658

    Figure Lengend Snippet: AKT is essential for the modulation of keratinocyte differentiation by HPV18 (A) Schematic showing the KGFR signalling pathway and the effects of the dominant active (DA) and negative (DN) AKT. HPV18 genome containing keratinocytes were infected with empty retrovirus, or with retroviruses encoding DA AKT (B) or DN AKT (C) . Cells were differentiated in high calcium media for 48 hours prior to lysis and analysed for expression of cytokeratin 1 (K1) to assess differentiation. Expression of the exogenous AKT was confirmed using an antibody against the HA epitope and GAPDH served as a loading control.

    Article Snippet: Retrovirus transduction pLNCX AKT vectors (Addgene 9005, 9006 [ ]) were transfected into HEK 293T cells with murine retrovirus envelope and GAG/polymerase plasmids (kindly provided by Professor Greg Towers, University College London) using PEI transfection reagent.

    Techniques: Infection, Lysis, Expressing

    NLRC3 regulates upstream signalling molecules within the PI3K–AKT–mTOR pathway a , Immunofluorescence staining of primary fibroblasts and frequency of co-localisation between mTOR and LAMP1 (n= > 150). b , Images and quantification of colon tumours in littermate Apc Min/+ and Apc Min/+ Nlrc3 −/− mice 40 days after treatment with vehicle or NVP-BEZ235. c , Immunohistochemical staining of colon tissues from mice treated as in b. d , Immunoprecipitation of the GFP tag in 239T cells transfected with a plasmid encoding GFP alone or NLRC3-GFP. e , Immunoblot of mouse colon tissues and densitometric quantification. f , Immunoblot of primary mouse fibroblasts transduced with a retroviral vector encoding GFP or NLRC3-GFP, with or without stimulation with IGF-1, and densitometric quantification. Scale bar, 10 µm ( a ), 200 µm ( c ). * P

    Journal: Nature

    Article Title: NLRC3 is an inhibitory sensor of PI3K–mTOR pathways in cancer

    doi: 10.1038/nature20597

    Figure Lengend Snippet: NLRC3 regulates upstream signalling molecules within the PI3K–AKT–mTOR pathway a , Immunofluorescence staining of primary fibroblasts and frequency of co-localisation between mTOR and LAMP1 (n= > 150). b , Images and quantification of colon tumours in littermate Apc Min/+ and Apc Min/+ Nlrc3 −/− mice 40 days after treatment with vehicle or NVP-BEZ235. c , Immunohistochemical staining of colon tissues from mice treated as in b. d , Immunoprecipitation of the GFP tag in 239T cells transfected with a plasmid encoding GFP alone or NLRC3-GFP. e , Immunoblot of mouse colon tissues and densitometric quantification. f , Immunoblot of primary mouse fibroblasts transduced with a retroviral vector encoding GFP or NLRC3-GFP, with or without stimulation with IGF-1, and densitometric quantification. Scale bar, 10 µm ( a ), 200 µm ( c ). * P

    Article Snippet: Retroviral transduction Human or mouse MSCV-NLRC3-IRES-GFP or MSCV-IRES-GFP was co-transfected with a retroviral packaging plasmids (pPAM-E and VSV-G) into HEK293T cells using Xfect Transfection Reagents (631318, Clontech Laboratories, Inc.).

    Techniques: Immunofluorescence, Staining, Mouse Assay, Immunohistochemistry, Immunoprecipitation, Transfection, Plasmid Preparation, Transduction

    Autophagic flux in virus-specific CD8 T cells is inversely correlated with cell proliferation status. ( a ) MSCV retroviral constructs showing the transgenes (GFP-LC3b and GFP-LC3b-G120A) introduced in P14 cells to probe for autophagy activity. ( b ) Flow cytometry plots of adoptively transferred P14 cells in spleens. P14 cells transduced with retroviral plasmid of either MIT-GFP-LC3b or MIT-G120A are positive for the congenic marker Thy1.1. Day 5, 8 and 30 p.i. splenocytes were used for the analysis. The percentage of GFP-negative cells out of the transduced P14 (Thy1.1 + ) cells from each group is highlighted in blue on the lower right corner of each plot and is summarized in ( c ). ( d ) Representative ImageStream ® data showing images of P14 cells from either GFP-LC3b or G120A groups on day 8 p.i.. Images showing GFP signals of GFP + MIT vector-transduced P14 cells (Ly5.1 + Thy1.1 + ). Summary graph of ImageStream ® analysis is plotted in ( e ) showing percentage of GFP + or Thy1.1 + P14 cells that exhibited more than one GFP punctum. Errors bars represent SEM. ( b ) and ( d ) are representative of three independent experiments, n≥3 mice in each group.

    Journal: Nature immunology

    Article Title: Autophagy is essential for effector CD8 T cell survival and memory formation

    doi: 10.1038/ni.3025

    Figure Lengend Snippet: Autophagic flux in virus-specific CD8 T cells is inversely correlated with cell proliferation status. ( a ) MSCV retroviral constructs showing the transgenes (GFP-LC3b and GFP-LC3b-G120A) introduced in P14 cells to probe for autophagy activity. ( b ) Flow cytometry plots of adoptively transferred P14 cells in spleens. P14 cells transduced with retroviral plasmid of either MIT-GFP-LC3b or MIT-G120A are positive for the congenic marker Thy1.1. Day 5, 8 and 30 p.i. splenocytes were used for the analysis. The percentage of GFP-negative cells out of the transduced P14 (Thy1.1 + ) cells from each group is highlighted in blue on the lower right corner of each plot and is summarized in ( c ). ( d ) Representative ImageStream ® data showing images of P14 cells from either GFP-LC3b or G120A groups on day 8 p.i.. Images showing GFP signals of GFP + MIT vector-transduced P14 cells (Ly5.1 + Thy1.1 + ). Summary graph of ImageStream ® analysis is plotted in ( e ) showing percentage of GFP + or Thy1.1 + P14 cells that exhibited more than one GFP punctum. Errors bars represent SEM. ( b ) and ( d ) are representative of three independent experiments, n≥3 mice in each group.

    Article Snippet: Retroviral transduction Over-expression of transgenes in P14 was carried out using MSCV-IRES-Thy1.1 retrovirus vector (kindly provided by A. Rao, La Jolla Institute for Allergy and Immunology, La Jolla, CA; Addgene plasmid 17442).

    Techniques: Construct, Activity Assay, Flow Cytometry, Cytometry, Transduction, Plasmid Preparation, Marker, Mouse Assay