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  • 87
    OriGene complementary dna encoding full length flag tagged tif 1 γ
    Complementary Dna Encoding Full Length Flag Tagged Tif 1 γ, supplied by OriGene, used in various techniques. Bioz Stars score: 87/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 87 stars, based on 7 article reviews
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    92
    Thermo Fisher myc tagged cdna
    Myc Tagged Cdna, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 24 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    OriGene ddk tagged mplscr1 cdna
    Ddk Tagged Mplscr1 Cdna, supplied by OriGene, used in various techniques. Bioz Stars score: 93/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    80
    Addgene inc flag tagged gfi1 cdna
    In vivo transfection of cerebellar cells with <t>Gfi1</t> / c-MYC and Otx2 / c-MYC induces Group 3 MB. a DAPI staining and GFP immunofluorescence of CD1 mouse brain section 3 months after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. b DAPI and GFP immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. c Hematoxylin and <t>Flag</t> (Gfi1) immunohistochemistry of CD1 mouse tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. d DAPI and Otx2 immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. e , f Hematoxylin and NPR3 immunohistochemistry of CD1 mice tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( e ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( f ). g , h GFP and GFAP immunofluorescence of tumors in CD1 mice after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( g ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( h ). i In vivo bioluminescent imaging of the upper body of a CD1 mouse transfected with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0, with a plasmid encoding firefly luciferase (pPBLuc). Imaging was performed weekly from the sixth week after transfection. j In vivo bioluminescent imaging of the lumbosacral region of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. The luminescence signal is expressed in photons/sec/mm2. k DAPI and immunofluorescence for pH3 of transversal section of the sacral portion of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l DAPI and immunofluorescence for GFP and pH3 of sacral transversal section of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l is the higher magnification of the white square in ( k ). Scale bars 1 mm in ( a , b ), 100 µm in ( c – h ), 500 µm in ( k ).
    Flag Tagged Gfi1 Cdna, supplied by Addgene inc, used in various techniques. Bioz Stars score: 80/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Genecopoeia myc tagged marf1 cdna
    In vivo transfection of cerebellar cells with <t>Gfi1</t> / c-MYC and Otx2 / c-MYC induces Group 3 MB. a DAPI staining and GFP immunofluorescence of CD1 mouse brain section 3 months after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. b DAPI and GFP immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. c Hematoxylin and <t>Flag</t> (Gfi1) immunohistochemistry of CD1 mouse tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. d DAPI and Otx2 immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. e , f Hematoxylin and NPR3 immunohistochemistry of CD1 mice tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( e ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( f ). g , h GFP and GFAP immunofluorescence of tumors in CD1 mice after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( g ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( h ). i In vivo bioluminescent imaging of the upper body of a CD1 mouse transfected with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0, with a plasmid encoding firefly luciferase (pPBLuc). Imaging was performed weekly from the sixth week after transfection. j In vivo bioluminescent imaging of the lumbosacral region of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. The luminescence signal is expressed in photons/sec/mm2. k DAPI and immunofluorescence for pH3 of transversal section of the sacral portion of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l DAPI and immunofluorescence for GFP and pH3 of sacral transversal section of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l is the higher magnification of the white square in ( k ). Scale bars 1 mm in ( a , b ), 100 µm in ( c – h ), 500 µm in ( k ).
    Myc Tagged Marf1 Cdna, supplied by Genecopoeia, 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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    93
    Genecopoeia er ha tag cdna
    In vivo transfection of cerebellar cells with <t>Gfi1</t> / c-MYC and Otx2 / c-MYC induces Group 3 MB. a DAPI staining and GFP immunofluorescence of CD1 mouse brain section 3 months after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. b DAPI and GFP immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. c Hematoxylin and <t>Flag</t> (Gfi1) immunohistochemistry of CD1 mouse tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. d DAPI and Otx2 immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. e , f Hematoxylin and NPR3 immunohistochemistry of CD1 mice tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( e ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( f ). g , h GFP and GFAP immunofluorescence of tumors in CD1 mice after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( g ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( h ). i In vivo bioluminescent imaging of the upper body of a CD1 mouse transfected with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0, with a plasmid encoding firefly luciferase (pPBLuc). Imaging was performed weekly from the sixth week after transfection. j In vivo bioluminescent imaging of the lumbosacral region of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. The luminescence signal is expressed in photons/sec/mm2. k DAPI and immunofluorescence for pH3 of transversal section of the sacral portion of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l DAPI and immunofluorescence for GFP and pH3 of sacral transversal section of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l is the higher magnification of the white square in ( k ). Scale bars 1 mm in ( a , b ), 100 µm in ( c – h ), 500 µm in ( k ).
    Er Ha Tag Cdna, supplied by Genecopoeia, 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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    93
    OriGene slitrk4 gfp tagged cdna
    In vivo transfection of cerebellar cells with <t>Gfi1</t> / c-MYC and Otx2 / c-MYC induces Group 3 MB. a DAPI staining and GFP immunofluorescence of CD1 mouse brain section 3 months after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. b DAPI and GFP immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. c Hematoxylin and <t>Flag</t> (Gfi1) immunohistochemistry of CD1 mouse tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. d DAPI and Otx2 immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. e , f Hematoxylin and NPR3 immunohistochemistry of CD1 mice tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( e ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( f ). g , h GFP and GFAP immunofluorescence of tumors in CD1 mice after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( g ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( h ). i In vivo bioluminescent imaging of the upper body of a CD1 mouse transfected with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0, with a plasmid encoding firefly luciferase (pPBLuc). Imaging was performed weekly from the sixth week after transfection. j In vivo bioluminescent imaging of the lumbosacral region of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. The luminescence signal is expressed in photons/sec/mm2. k DAPI and immunofluorescence for pH3 of transversal section of the sacral portion of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l DAPI and immunofluorescence for GFP and pH3 of sacral transversal section of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l is the higher magnification of the white square in ( k ). Scale bars 1 mm in ( a , b ), 100 µm in ( c – h ), 500 µm in ( k ).
    Slitrk4 Gfp Tagged Cdna, supplied by OriGene, used in various techniques. Bioz Stars score: 93/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    Addgene inc flag tagged p21 cdna
    In vivo transfection of cerebellar cells with <t>Gfi1</t> / c-MYC and Otx2 / c-MYC induces Group 3 MB. a DAPI staining and GFP immunofluorescence of CD1 mouse brain section 3 months after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. b DAPI and GFP immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. c Hematoxylin and <t>Flag</t> (Gfi1) immunohistochemistry of CD1 mouse tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. d DAPI and Otx2 immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. e , f Hematoxylin and NPR3 immunohistochemistry of CD1 mice tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( e ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( f ). g , h GFP and GFAP immunofluorescence of tumors in CD1 mice after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( g ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( h ). i In vivo bioluminescent imaging of the upper body of a CD1 mouse transfected with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0, with a plasmid encoding firefly luciferase (pPBLuc). Imaging was performed weekly from the sixth week after transfection. j In vivo bioluminescent imaging of the lumbosacral region of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. The luminescence signal is expressed in photons/sec/mm2. k DAPI and immunofluorescence for pH3 of transversal section of the sacral portion of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l DAPI and immunofluorescence for GFP and pH3 of sacral transversal section of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l is the higher magnification of the white square in ( k ). Scale bars 1 mm in ( a , b ), 100 µm in ( c – h ), 500 µm in ( k ).
    Flag Tagged P21 Cdna, supplied by Addgene inc, used in various techniques. Bioz Stars score: 86/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Genecopoeia runx2 ha tag cdna
    In vivo transfection of cerebellar cells with <t>Gfi1</t> / c-MYC and Otx2 / c-MYC induces Group 3 MB. a DAPI staining and GFP immunofluorescence of CD1 mouse brain section 3 months after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. b DAPI and GFP immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. c Hematoxylin and <t>Flag</t> (Gfi1) immunohistochemistry of CD1 mouse tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. d DAPI and Otx2 immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. e , f Hematoxylin and NPR3 immunohistochemistry of CD1 mice tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( e ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( f ). g , h GFP and GFAP immunofluorescence of tumors in CD1 mice after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( g ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( h ). i In vivo bioluminescent imaging of the upper body of a CD1 mouse transfected with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0, with a plasmid encoding firefly luciferase (pPBLuc). Imaging was performed weekly from the sixth week after transfection. j In vivo bioluminescent imaging of the lumbosacral region of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. The luminescence signal is expressed in photons/sec/mm2. k DAPI and immunofluorescence for pH3 of transversal section of the sacral portion of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l DAPI and immunofluorescence for GFP and pH3 of sacral transversal section of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l is the higher magnification of the white square in ( k ). Scale bars 1 mm in ( a , b ), 100 µm in ( c – h ), 500 µm in ( k ).
    Runx2 Ha Tag Cdna, supplied by Genecopoeia, used in various techniques. Bioz Stars score: 92/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    OriGene myc ddk tagged zfp407 cdna
    In vivo transfection of cerebellar cells with <t>Gfi1</t> / c-MYC and Otx2 / c-MYC induces Group 3 MB. a DAPI staining and GFP immunofluorescence of CD1 mouse brain section 3 months after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. b DAPI and GFP immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. c Hematoxylin and <t>Flag</t> (Gfi1) immunohistochemistry of CD1 mouse tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. d DAPI and Otx2 immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. e , f Hematoxylin and NPR3 immunohistochemistry of CD1 mice tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( e ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( f ). g , h GFP and GFAP immunofluorescence of tumors in CD1 mice after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( g ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( h ). i In vivo bioluminescent imaging of the upper body of a CD1 mouse transfected with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0, with a plasmid encoding firefly luciferase (pPBLuc). Imaging was performed weekly from the sixth week after transfection. j In vivo bioluminescent imaging of the lumbosacral region of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. The luminescence signal is expressed in photons/sec/mm2. k DAPI and immunofluorescence for pH3 of transversal section of the sacral portion of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l DAPI and immunofluorescence for GFP and pH3 of sacral transversal section of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l is the higher magnification of the white square in ( k ). Scale bars 1 mm in ( a , b ), 100 µm in ( c – h ), 500 µm in ( k ).
    Myc Ddk Tagged Zfp407 Cdna, supplied by OriGene, 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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    94
    Addgene inc flag tagged lkb1 cdna
    Genetic experiments establish an essential <t>LKB1-SIK3-HDAC4</t> pathway in AML (A) SIK3 mutants used in this study. (B) Western blot in the cells transduced with empty vector or the indicated SIK3 <t>cDNA.</t> (C and D) Competition-based proliferation assays in the cells transduced with empty vector or the indicated SIK3 cDNA and infected with the indicated GFP-linked sgRNA.. (E) Chemical structure of HG-9-91-01. (F – H) Relative growth of the indicated cells, harboring empty vector, SIK3, or SIK3 T142Q cDNA, upon HG-9-91-01 treatment. Normalized relative luminescence unit (RLU) was shown after 3 days culture with DMSO (0.1%) or HG-9-91-01 at the indicated concentrations. The mean ± SEM (n=3) and four-parameter dose-response curves are plotted. (I) Competition-based proliferation assays in the cells co-infected with GFP-linked sgRNA and mCherry-linked sgRNA. The percentage of double mCherry+/GFP+ cells is shown. (J) Western blot in the cells infected with the indicated sgRNA. (K) Western blot in the cells harboring empty vector, SIK3, or SIK3 T142Q cDNA, followed by treatment with DMSO (0.1%) or 100 nM HG-9-91-01 for 2 hrs. All bar graphs represent the mean ± SEM (n=3). Neg1, Neg2: negative controls. .
    Flag Tagged Lkb1 Cdna, supplied by Addgene inc, used in various techniques. Bioz Stars score: 94/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    OriGene myc ddk tagged brca1 cdna
    Genetic experiments establish an essential <t>LKB1-SIK3-HDAC4</t> pathway in AML (A) SIK3 mutants used in this study. (B) Western blot in the cells transduced with empty vector or the indicated SIK3 <t>cDNA.</t> (C and D) Competition-based proliferation assays in the cells transduced with empty vector or the indicated SIK3 cDNA and infected with the indicated GFP-linked sgRNA.. (E) Chemical structure of HG-9-91-01. (F – H) Relative growth of the indicated cells, harboring empty vector, SIK3, or SIK3 T142Q cDNA, upon HG-9-91-01 treatment. Normalized relative luminescence unit (RLU) was shown after 3 days culture with DMSO (0.1%) or HG-9-91-01 at the indicated concentrations. The mean ± SEM (n=3) and four-parameter dose-response curves are plotted. (I) Competition-based proliferation assays in the cells co-infected with GFP-linked sgRNA and mCherry-linked sgRNA. The percentage of double mCherry+/GFP+ cells is shown. (J) Western blot in the cells infected with the indicated sgRNA. (K) Western blot in the cells harboring empty vector, SIK3, or SIK3 T142Q cDNA, followed by treatment with DMSO (0.1%) or 100 nM HG-9-91-01 for 2 hrs. All bar graphs represent the mean ± SEM (n=3). Neg1, Neg2: negative controls. .
    Myc Ddk Tagged Brca1 Cdna, supplied by OriGene, used in various techniques. Bioz Stars score: 95/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Genecopoeia egfp tagged cdna constructs
    Genetic experiments establish an essential <t>LKB1-SIK3-HDAC4</t> pathway in AML (A) SIK3 mutants used in this study. (B) Western blot in the cells transduced with empty vector or the indicated SIK3 <t>cDNA.</t> (C and D) Competition-based proliferation assays in the cells transduced with empty vector or the indicated SIK3 cDNA and infected with the indicated GFP-linked sgRNA.. (E) Chemical structure of HG-9-91-01. (F – H) Relative growth of the indicated cells, harboring empty vector, SIK3, or SIK3 T142Q cDNA, upon HG-9-91-01 treatment. Normalized relative luminescence unit (RLU) was shown after 3 days culture with DMSO (0.1%) or HG-9-91-01 at the indicated concentrations. The mean ± SEM (n=3) and four-parameter dose-response curves are plotted. (I) Competition-based proliferation assays in the cells co-infected with GFP-linked sgRNA and mCherry-linked sgRNA. The percentage of double mCherry+/GFP+ cells is shown. (J) Western blot in the cells infected with the indicated sgRNA. (K) Western blot in the cells harboring empty vector, SIK3, or SIK3 T142Q cDNA, followed by treatment with DMSO (0.1%) or 100 nM HG-9-91-01 for 2 hrs. All bar graphs represent the mean ± SEM (n=3). Neg1, Neg2: negative controls. .
    Egfp Tagged Cdna Constructs, supplied by Genecopoeia, used in various techniques. Bioz Stars score: 90/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    OriGene gfp tagged murine ido cdna
    <t>IDO</t> impairs antitumor effects of anti-CTLA4 in the context of B16 tumors engineered to overexpress IDO. (A) Expression of <t>GFP</t> (black line) or IDO/GFP (gray) in B16F10 cells determined by flow cytometry. WT B16F10 cells were used for comparison (dotted line). (B) In vitro growth rate of GFP-B16F10 and IDO/GFP-B16F10 cells. (C) Mean growth of GFP-B16F10 and IDO/GFP-B16F10 tumors in naive and irradiated (450 cGy) mice. (D) Tumor-free survival curves for C57BL/6 mice challenged with GFP-B16F10 or IDO/GFP-B16F10 tumor cells i.d. and treated with anti–CTLA-4 or anti-CLTA-4/1MT. Data represent cumulative results from two independent experiments with five mice/group. Statistical significance was determined with Log-Rank test (*, P
    Gfp Tagged Murine Ido Cdna, supplied by OriGene, 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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    85
    Illumina Inc cdna 5 tags
    <t>IDO</t> impairs antitumor effects of anti-CTLA4 in the context of B16 tumors engineered to overexpress IDO. (A) Expression of <t>GFP</t> (black line) or IDO/GFP (gray) in B16F10 cells determined by flow cytometry. WT B16F10 cells were used for comparison (dotted line). (B) In vitro growth rate of GFP-B16F10 and IDO/GFP-B16F10 cells. (C) Mean growth of GFP-B16F10 and IDO/GFP-B16F10 tumors in naive and irradiated (450 cGy) mice. (D) Tumor-free survival curves for C57BL/6 mice challenged with GFP-B16F10 or IDO/GFP-B16F10 tumor cells i.d. and treated with anti–CTLA-4 or anti-CLTA-4/1MT. Data represent cumulative results from two independent experiments with five mice/group. Statistical significance was determined with Log-Rank test (*, P
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    Illumina Inc illumina cdna tags
    <t>IDO</t> impairs antitumor effects of anti-CTLA4 in the context of B16 tumors engineered to overexpress IDO. (A) Expression of <t>GFP</t> (black line) or IDO/GFP (gray) in B16F10 cells determined by flow cytometry. WT B16F10 cells were used for comparison (dotted line). (B) In vitro growth rate of GFP-B16F10 and IDO/GFP-B16F10 cells. (C) Mean growth of GFP-B16F10 and IDO/GFP-B16F10 tumors in naive and irradiated (450 cGy) mice. (D) Tumor-free survival curves for C57BL/6 mice challenged with GFP-B16F10 or IDO/GFP-B16F10 tumor cells i.d. and treated with anti–CTLA-4 or anti-CLTA-4/1MT. Data represent cumulative results from two independent experiments with five mice/group. Statistical significance was determined with Log-Rank test (*, P
    Illumina Cdna Tags, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 84/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    OriGene gfp tagged gfap cdna
    Characterization of the <t>GFAP</t> c.1289G > A/p.R430H mutation. A : Schematic representation of the exonic structure of different GFAP isoforms. Dotted lines indicate the termination codons. The arrows indicate the position of the c.1289G > A variant (Note that in GFAP-κ the c.1289G > A mutation is part of the 3′-UTR). B : Electropherograms of GFAP exon 7A region containing c.1289G > A variant, in patients 1 and 2 (Pt1, Pt2) and in their mother (I-2). C : The histogram displays the percentages of cells transfected with <t>GFP-GFAP-ϵ</t> wt (green bars) or GFP-GFAP-ϵ R430H (purple bars), classified in filamentous pattern (F), cytoplasmic aggregates on a filamentous pattern (F + A), cytoplasmic aggregates with no filamentous pattern (A). Scale bars represent 15 μm. A total of 324 cells for GFP-GFAP-ϵ wt and 285 for GFP-GFAP-ϵ R430H , from 3 independent experiments, were blindly analyzed by two different operators. ANOVA test for interaction p = 0.001.
    Gfp Tagged Gfap Cdna, supplied by OriGene, used in various techniques. Bioz Stars score: 86/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Roche flx cdna tags
    Characterization of the <t>GFAP</t> c.1289G > A/p.R430H mutation. A : Schematic representation of the exonic structure of different GFAP isoforms. Dotted lines indicate the termination codons. The arrows indicate the position of the c.1289G > A variant (Note that in GFAP-κ the c.1289G > A mutation is part of the 3′-UTR). B : Electropherograms of GFAP exon 7A region containing c.1289G > A variant, in patients 1 and 2 (Pt1, Pt2) and in their mother (I-2). C : The histogram displays the percentages of cells transfected with <t>GFP-GFAP-ϵ</t> wt (green bars) or GFP-GFAP-ϵ R430H (purple bars), classified in filamentous pattern (F), cytoplasmic aggregates on a filamentous pattern (F + A), cytoplasmic aggregates with no filamentous pattern (A). Scale bars represent 15 μm. A total of 324 cells for GFP-GFAP-ϵ wt and 285 for GFP-GFAP-ϵ R430H , from 3 independent experiments, were blindly analyzed by two different operators. ANOVA test for interaction p = 0.001.
    Flx Cdna Tags, supplied by Roche, 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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    TaKaRa adaptor tagged cdna
    Characterization of the <t>GFAP</t> c.1289G > A/p.R430H mutation. A : Schematic representation of the exonic structure of different GFAP isoforms. Dotted lines indicate the termination codons. The arrows indicate the position of the c.1289G > A variant (Note that in GFAP-κ the c.1289G > A mutation is part of the 3′-UTR). B : Electropherograms of GFAP exon 7A region containing c.1289G > A variant, in patients 1 and 2 (Pt1, Pt2) and in their mother (I-2). C : The histogram displays the percentages of cells transfected with <t>GFP-GFAP-ϵ</t> wt (green bars) or GFP-GFAP-ϵ R430H (purple bars), classified in filamentous pattern (F), cytoplasmic aggregates on a filamentous pattern (F + A), cytoplasmic aggregates with no filamentous pattern (A). Scale bars represent 15 μm. A total of 324 cells for GFP-GFAP-ϵ wt and 285 for GFP-GFAP-ϵ R430H , from 3 independent experiments, were blindly analyzed by two different operators. ANOVA test for interaction p = 0.001.
    Adaptor Tagged Cdna, supplied by TaKaRa, used in various techniques. Bioz Stars score: 84/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Sino Biological vegfr 3 flag tagged cdna
    uPARAP forms a complex with VEGFR-2 and <t>VEGFR-3</t> and restricts VEGFR-2/VEGFR-3 heterodimerisation. In situ PLA in LECs ( a , b , e – i ) or PAECs ( d ) stimulated or not for 5 (T5) and 30 min (T30) with VEGF-C ( a , b , e – g ). a , b Interaction between uPARAP and VEGFR-2 ( a ) or VEGFR-3 ( b ). Results are those of one representative assay out of 3. Data represent mean (biological duplicates, 20 images analysed for quantification) ± SEM. Bars = 10 µm. c Immunoprecipitation (IP) of uPARAP at the indicated time point of VEGF-C stimulation. Western blots were revealed with an antibody (IB) against VEGFR-2, VEGFR-3 (upper panel) or uPARAP (lower panel). GAPDH was used as loading control. d In situ PLA detection of uPARAP interaction with VEGFR-2 or VEGFR-3 under basal conditions, in PAECs expressing VEGFR-2 (PAEC-VEGFR-2), VEGFR-3 (PAEC-VEGFR-3) or no VEGFR (PAEC). Bars = 10 µm. e , f In situ PLA detection of VEGFR-2/VEGFR-3 heterodimerisation in control LECs (Ctr), LECs deficient for uPARAP expression (siU1 and siU2) ( e ), or LECs overexpressing uPARAP (uPARAP <t>cDNA)</t> ( f ). g VEGFR-3 homodimerisation in LECs transfected with a VEGFR-3-Flag-tagged cDNA. Data correspond to the mean (biological triplicates, 60 images analysed for quantification, > 100 cells analysed per condition) ± SEM ( e – g ). h In situ PLA detection of uPARAP/VEGFR-2 and uPARAP/VEGFR-3 complexes in LECs migrating in a VEGF-C gradient. Bars = 20 µm. i In situ PLA detection of VEGFR-2/VEGFR-3 heterodimers in control LECs (Ctr) and uPARAP silenced LECs (siU) migrating in a VEGF-C gradient. The left histogram corresponds to the mean number of dots per cell. The right histogram represents PLA signal localisation with respect to the nucleus (one representative experiment out of 3 with 25 cells analysed per condition in each experiment). Bars = 20 µm. Statistical analyses were performed using a non-parametric Mann–Whitney test. i Chi-square test was used. * P
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    OriGene flag tagged pdha1 cdna sequence
    uPARAP forms a complex with VEGFR-2 and <t>VEGFR-3</t> and restricts VEGFR-2/VEGFR-3 heterodimerisation. In situ PLA in LECs ( a , b , e – i ) or PAECs ( d ) stimulated or not for 5 (T5) and 30 min (T30) with VEGF-C ( a , b , e – g ). a , b Interaction between uPARAP and VEGFR-2 ( a ) or VEGFR-3 ( b ). Results are those of one representative assay out of 3. Data represent mean (biological duplicates, 20 images analysed for quantification) ± SEM. Bars = 10 µm. c Immunoprecipitation (IP) of uPARAP at the indicated time point of VEGF-C stimulation. Western blots were revealed with an antibody (IB) against VEGFR-2, VEGFR-3 (upper panel) or uPARAP (lower panel). GAPDH was used as loading control. d In situ PLA detection of uPARAP interaction with VEGFR-2 or VEGFR-3 under basal conditions, in PAECs expressing VEGFR-2 (PAEC-VEGFR-2), VEGFR-3 (PAEC-VEGFR-3) or no VEGFR (PAEC). Bars = 10 µm. e , f In situ PLA detection of VEGFR-2/VEGFR-3 heterodimerisation in control LECs (Ctr), LECs deficient for uPARAP expression (siU1 and siU2) ( e ), or LECs overexpressing uPARAP (uPARAP <t>cDNA)</t> ( f ). g VEGFR-3 homodimerisation in LECs transfected with a VEGFR-3-Flag-tagged cDNA. Data correspond to the mean (biological triplicates, 60 images analysed for quantification, > 100 cells analysed per condition) ± SEM ( e – g ). h In situ PLA detection of uPARAP/VEGFR-2 and uPARAP/VEGFR-3 complexes in LECs migrating in a VEGF-C gradient. Bars = 20 µm. i In situ PLA detection of VEGFR-2/VEGFR-3 heterodimers in control LECs (Ctr) and uPARAP silenced LECs (siU) migrating in a VEGF-C gradient. The left histogram corresponds to the mean number of dots per cell. The right histogram represents PLA signal localisation with respect to the nucleus (one representative experiment out of 3 with 25 cells analysed per condition in each experiment). Bars = 20 µm. Statistical analyses were performed using a non-parametric Mann–Whitney test. i Chi-square test was used. * P
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    OriGene myc ddk tagged ptgr1 cdna
    uPARAP forms a complex with VEGFR-2 and <t>VEGFR-3</t> and restricts VEGFR-2/VEGFR-3 heterodimerisation. In situ PLA in LECs ( a , b , e – i ) or PAECs ( d ) stimulated or not for 5 (T5) and 30 min (T30) with VEGF-C ( a , b , e – g ). a , b Interaction between uPARAP and VEGFR-2 ( a ) or VEGFR-3 ( b ). Results are those of one representative assay out of 3. Data represent mean (biological duplicates, 20 images analysed for quantification) ± SEM. Bars = 10 µm. c Immunoprecipitation (IP) of uPARAP at the indicated time point of VEGF-C stimulation. Western blots were revealed with an antibody (IB) against VEGFR-2, VEGFR-3 (upper panel) or uPARAP (lower panel). GAPDH was used as loading control. d In situ PLA detection of uPARAP interaction with VEGFR-2 or VEGFR-3 under basal conditions, in PAECs expressing VEGFR-2 (PAEC-VEGFR-2), VEGFR-3 (PAEC-VEGFR-3) or no VEGFR (PAEC). Bars = 10 µm. e , f In situ PLA detection of VEGFR-2/VEGFR-3 heterodimerisation in control LECs (Ctr), LECs deficient for uPARAP expression (siU1 and siU2) ( e ), or LECs overexpressing uPARAP (uPARAP <t>cDNA)</t> ( f ). g VEGFR-3 homodimerisation in LECs transfected with a VEGFR-3-Flag-tagged cDNA. Data correspond to the mean (biological triplicates, 60 images analysed for quantification, > 100 cells analysed per condition) ± SEM ( e – g ). h In situ PLA detection of uPARAP/VEGFR-2 and uPARAP/VEGFR-3 complexes in LECs migrating in a VEGF-C gradient. Bars = 20 µm. i In situ PLA detection of VEGFR-2/VEGFR-3 heterodimers in control LECs (Ctr) and uPARAP silenced LECs (siU) migrating in a VEGF-C gradient. The left histogram corresponds to the mean number of dots per cell. The right histogram represents PLA signal localisation with respect to the nucleus (one representative experiment out of 3 with 25 cells analysed per condition in each experiment). Bars = 20 µm. Statistical analyses were performed using a non-parametric Mann–Whitney test. i Chi-square test was used. * P
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    OriGene gfp tagged foxo1 cdna
    PTEN/NLK synthetic lethality is abrogated by <t>FOXO1</t> silencing. ( A ) Survival analysis of HCT116 cells transfected with NLK and/or FOXO1 siRNA. HCT116 PTEN −/− and HCT116 PTEN +/+ cells were transfected with siRNA targeting NLK and FOXO1 as shown and surviving fractions determined after five days. The p value (*) was calculated using Student's t test. ( B ) Nuclear localisation of FOXO1 is enhanced in PTEN deficient cells upon NLK silencing. HCT116 isogenic cells were co-transfected with <t>GFP-tagged</t> FOXO1 <t>cDNA</t> in addition to control (non targeting) siRNA (siCON) or NLK siRNA. Two days later cells were fixed and stained with DAPI. Green signal represents GFP-FOXO1 and blue signal represents nuclear DAPI (nuclear) staining. Arrows indicate cells with nuclear localisation of FOXO1. ( C ) Senescence is increased by NLK siRNA in PTEN deficient cells. Bar chart of relative relative senescence levels caused by NLK silencing are shown. HCT116-derived PTEN isogenic cell lines were reversed transfected with a pool of two validated siRNAs against NLK, as well as siCON pool#2 (Dharmacon) as non-targeting control, using RNAiMAX (Invitrogen). Seven days after transfection cells were fixed and incubated overnight at 37°C in a solution containing X-gal. ( D ) Representative images for β-Galactosidase staining of PTEN deficient cells. Blue staining indicates β-Galactosidase.
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    OriGene myc tagged human kdm4d cdna
    PTEN/NLK synthetic lethality is abrogated by <t>FOXO1</t> silencing. ( A ) Survival analysis of HCT116 cells transfected with NLK and/or FOXO1 siRNA. HCT116 PTEN −/− and HCT116 PTEN +/+ cells were transfected with siRNA targeting NLK and FOXO1 as shown and surviving fractions determined after five days. The p value (*) was calculated using Student's t test. ( B ) Nuclear localisation of FOXO1 is enhanced in PTEN deficient cells upon NLK silencing. HCT116 isogenic cells were co-transfected with <t>GFP-tagged</t> FOXO1 <t>cDNA</t> in addition to control (non targeting) siRNA (siCON) or NLK siRNA. Two days later cells were fixed and stained with DAPI. Green signal represents GFP-FOXO1 and blue signal represents nuclear DAPI (nuclear) staining. Arrows indicate cells with nuclear localisation of FOXO1. ( C ) Senescence is increased by NLK siRNA in PTEN deficient cells. Bar chart of relative relative senescence levels caused by NLK silencing are shown. HCT116-derived PTEN isogenic cell lines were reversed transfected with a pool of two validated siRNAs against NLK, as well as siCON pool#2 (Dharmacon) as non-targeting control, using RNAiMAX (Invitrogen). Seven days after transfection cells were fixed and incubated overnight at 37°C in a solution containing X-gal. ( D ) Representative images for β-Galactosidase staining of PTEN deficient cells. Blue staining indicates β-Galactosidase.
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    Addgene inc flag tagged human yod1 cdna
    PTEN/NLK synthetic lethality is abrogated by <t>FOXO1</t> silencing. ( A ) Survival analysis of HCT116 cells transfected with NLK and/or FOXO1 siRNA. HCT116 PTEN −/− and HCT116 PTEN +/+ cells were transfected with siRNA targeting NLK and FOXO1 as shown and surviving fractions determined after five days. The p value (*) was calculated using Student's t test. ( B ) Nuclear localisation of FOXO1 is enhanced in PTEN deficient cells upon NLK silencing. HCT116 isogenic cells were co-transfected with <t>GFP-tagged</t> FOXO1 <t>cDNA</t> in addition to control (non targeting) siRNA (siCON) or NLK siRNA. Two days later cells were fixed and stained with DAPI. Green signal represents GFP-FOXO1 and blue signal represents nuclear DAPI (nuclear) staining. Arrows indicate cells with nuclear localisation of FOXO1. ( C ) Senescence is increased by NLK siRNA in PTEN deficient cells. Bar chart of relative relative senescence levels caused by NLK silencing are shown. HCT116-derived PTEN isogenic cell lines were reversed transfected with a pool of two validated siRNAs against NLK, as well as siCON pool#2 (Dharmacon) as non-targeting control, using RNAiMAX (Invitrogen). Seven days after transfection cells were fixed and incubated overnight at 37°C in a solution containing X-gal. ( D ) Representative images for β-Galactosidase staining of PTEN deficient cells. Blue staining indicates β-Galactosidase.
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    Thermo Fisher human cdna expressed sequence tags
    PTEN/NLK synthetic lethality is abrogated by <t>FOXO1</t> silencing. ( A ) Survival analysis of HCT116 cells transfected with NLK and/or FOXO1 siRNA. HCT116 PTEN −/− and HCT116 PTEN +/+ cells were transfected with siRNA targeting NLK and FOXO1 as shown and surviving fractions determined after five days. The p value (*) was calculated using Student's t test. ( B ) Nuclear localisation of FOXO1 is enhanced in PTEN deficient cells upon NLK silencing. HCT116 isogenic cells were co-transfected with <t>GFP-tagged</t> FOXO1 <t>cDNA</t> in addition to control (non targeting) siRNA (siCON) or NLK siRNA. Two days later cells were fixed and stained with DAPI. Green signal represents GFP-FOXO1 and blue signal represents nuclear DAPI (nuclear) staining. Arrows indicate cells with nuclear localisation of FOXO1. ( C ) Senescence is increased by NLK siRNA in PTEN deficient cells. Bar chart of relative relative senescence levels caused by NLK silencing are shown. HCT116-derived PTEN isogenic cell lines were reversed transfected with a pool of two validated siRNAs against NLK, as well as siCON pool#2 (Dharmacon) as non-targeting control, using RNAiMAX (Invitrogen). Seven days after transfection cells were fixed and incubated overnight at 37°C in a solution containing X-gal. ( D ) Representative images for β-Galactosidase staining of PTEN deficient cells. Blue staining indicates β-Galactosidase.
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    Illumina Inc cdna sequence tags
    PTEN/NLK synthetic lethality is abrogated by <t>FOXO1</t> silencing. ( A ) Survival analysis of HCT116 cells transfected with NLK and/or FOXO1 siRNA. HCT116 PTEN −/− and HCT116 PTEN +/+ cells were transfected with siRNA targeting NLK and FOXO1 as shown and surviving fractions determined after five days. The p value (*) was calculated using Student's t test. ( B ) Nuclear localisation of FOXO1 is enhanced in PTEN deficient cells upon NLK silencing. HCT116 isogenic cells were co-transfected with <t>GFP-tagged</t> FOXO1 <t>cDNA</t> in addition to control (non targeting) siRNA (siCON) or NLK siRNA. Two days later cells were fixed and stained with DAPI. Green signal represents GFP-FOXO1 and blue signal represents nuclear DAPI (nuclear) staining. Arrows indicate cells with nuclear localisation of FOXO1. ( C ) Senescence is increased by NLK siRNA in PTEN deficient cells. Bar chart of relative relative senescence levels caused by NLK silencing are shown. HCT116-derived PTEN isogenic cell lines were reversed transfected with a pool of two validated siRNAs against NLK, as well as siCON pool#2 (Dharmacon) as non-targeting control, using RNAiMAX (Invitrogen). Seven days after transfection cells were fixed and incubated overnight at 37°C in a solution containing X-gal. ( D ) Representative images for β-Galactosidase staining of PTEN deficient cells. Blue staining indicates β-Galactosidase.
    Cdna Sequence Tags, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 87/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    CapitalBio Corporation cdna amplification tag kit
    PTEN/NLK synthetic lethality is abrogated by <t>FOXO1</t> silencing. ( A ) Survival analysis of HCT116 cells transfected with NLK and/or FOXO1 siRNA. HCT116 PTEN −/− and HCT116 PTEN +/+ cells were transfected with siRNA targeting NLK and FOXO1 as shown and surviving fractions determined after five days. The p value (*) was calculated using Student's t test. ( B ) Nuclear localisation of FOXO1 is enhanced in PTEN deficient cells upon NLK silencing. HCT116 isogenic cells were co-transfected with <t>GFP-tagged</t> FOXO1 <t>cDNA</t> in addition to control (non targeting) siRNA (siCON) or NLK siRNA. Two days later cells were fixed and stained with DAPI. Green signal represents GFP-FOXO1 and blue signal represents nuclear DAPI (nuclear) staining. Arrows indicate cells with nuclear localisation of FOXO1. ( C ) Senescence is increased by NLK siRNA in PTEN deficient cells. Bar chart of relative relative senescence levels caused by NLK silencing are shown. HCT116-derived PTEN isogenic cell lines were reversed transfected with a pool of two validated siRNAs against NLK, as well as siCON pool#2 (Dharmacon) as non-targeting control, using RNAiMAX (Invitrogen). Seven days after transfection cells were fixed and incubated overnight at 37°C in a solution containing X-gal. ( D ) Representative images for β-Galactosidase staining of PTEN deficient cells. Blue staining indicates β-Galactosidase.
    Cdna Amplification Tag Kit, supplied by CapitalBio Corporation, used in various techniques. Bioz Stars score: 86/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cassini egfp tagged cassini cdna
    Primer integrity and specificity analysis. ( A ) 2% agarose gel electrophoresis of control 8093 RT/PCR products using gapdh , 18 s rRNA , and <t>cassini</t> shows a single band for each target gene. ( B ) Real time RT/PCR on RNA isolated from non-transfected MEFs (control), MEFs transfected with an <t>EGFP-</t> cassini construct or with empty EGFP-C1 plasmid. Values (mean ± SD of triplicate real-time PCR) are expressed as percent of gapdh . ( C, D ) Derivative dissociation curves from real-time RT/PCR on mouse ( C ) and human ( D ) RNA confirm single product amplification. The cassini amplicon shows a single peak for both murine and human RNA with a T m of ~76°C. Gapdh primers peak at T m ~85.5°C. ***p
    Egfp Tagged Cassini Cdna, supplied by Cassini, used in various techniques. Bioz Stars score: 86/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Addgene inc flag tagged hdac6 cdna
    Increased intracellular sodium inhibits <t>HDAC6</t> activity. a DAOY cells were incubated with increasing concentrations of ouabain for three hours and then lysed for an in vitro HDAC activity assay as described in Materials and Methods. b DAOY cells were incubated with indicated concentrations of ouabain and the protein levels of the indicated HDAC isoforms were determined by immunoblotting. Actin immunoblot ensures equal loading. c After incubation with indicated concentrations of gramicidin A, HDAC6 was immunoprecipitated with a specific antibody and then subjected to an in vitro HDAC activity assay using a fluorescent substrate. d DAOY cells were transiently transfected with Flag-HDAC6 <t>cDNA.</t> The cells were incubated with the indicated concentrations of gramicidin A followed by immunoprecipitation of HDAC6. The immunoprecipitates were washed extensively and then incubated with polymerized microtubules to measure deacetylase activity. The reaction was terminated by adding lysis buffer and the samples were processed for immunoblotting with indicated antibodies. e Immunoblot for HDAC6 of stable clones of DAOY cells expressing HDAC6 shRNA. Control cells were transfected with vector only and processed in parallel. A GAPDH blot ensures that equal amounts of protein were used for analysis. f The HDAC6 knockdown cells were incubated with either High Na + or Low Na + buffer and the trafficking patterns of EGF-positive vesicles were analyzed as described in figure legend 3. *, P
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    90
    OriGene myc ddk tagged cdna
    Increased intracellular sodium inhibits <t>HDAC6</t> activity. a DAOY cells were incubated with increasing concentrations of ouabain for three hours and then lysed for an in vitro HDAC activity assay as described in Materials and Methods. b DAOY cells were incubated with indicated concentrations of ouabain and the protein levels of the indicated HDAC isoforms were determined by immunoblotting. Actin immunoblot ensures equal loading. c After incubation with indicated concentrations of gramicidin A, HDAC6 was immunoprecipitated with a specific antibody and then subjected to an in vitro HDAC activity assay using a fluorescent substrate. d DAOY cells were transiently transfected with Flag-HDAC6 <t>cDNA.</t> The cells were incubated with the indicated concentrations of gramicidin A followed by immunoprecipitation of HDAC6. The immunoprecipitates were washed extensively and then incubated with polymerized microtubules to measure deacetylase activity. The reaction was terminated by adding lysis buffer and the samples were processed for immunoblotting with indicated antibodies. e Immunoblot for HDAC6 of stable clones of DAOY cells expressing HDAC6 shRNA. Control cells were transfected with vector only and processed in parallel. A GAPDH blot ensures that equal amounts of protein were used for analysis. f The HDAC6 knockdown cells were incubated with either High Na + or Low Na + buffer and the trafficking patterns of EGF-positive vesicles were analyzed as described in figure legend 3. *, P
    Myc Ddk Tagged Cdna, supplied by OriGene, used in various techniques. Bioz Stars score: 90/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 90 stars, based on 18 article reviews
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    86
    OriGene flag tagged mouse gapdh cdna
    DDC-induced ROS formation is associated with a depletion of energy metabolism and antioxidant enzymes and is regulated by <t>GAPDH.</t> (A) Hepatocytes from C3H and C57BL mice were left untreated (−) or treated with either 0.1% DMSO vehicle (0) or the indicated concentrations of DDC for 48 h. Equal protein amounts (NP-40 lysates) were analyzed for expression of the indicated proteins. Samples from each strain were analyzed on separate gels. PDIA4 levels were unaltered (loading control). (B) C57BL hepatocytes were transfected with Control or GAPDH siRNA for 24 h and were then cultured for an additional 24 h in the presence of 0.025% DMSO vehicle (0) or 100 µM DDC or were left untreated (−). The NP-40 cell lysates were analyzed for the expression of the various proteins indicated. (C) Overexpression of Flag-tagged mouse GAPDH in isolated hepatocytes. NP-40 lysates were analyzed for expression of Flag-tagged GAPDH and total GAPDH. The Coomassie stain is included as a loading control. (D) Flag-GAPDH expression in C57BL hepatocytes, as determined by immunostaining with a mouse anti-Flag antibody (representing overexpressed GAPDH; green) and a rabbit anti-GAPDH antibody (representing total GAPDH; red). (E) C57BL hepatocytes were mock transfected (Control) or transfected with GAPDH siRNA or Flag-GAPDH mouse <t>cDNA</t> for 24 h and were then treated with 100 µM DDC for an additional 24 h. Representative images of ROS signal (green) with DAPI nuclear counterstain (blue) are shown. ROS levels (quantified as described in Materials and methods) exhibited a 2.2-fold increase after GAPDH knockdown and a 10-fold decrease after GAPDH overexpression relative to control. Bars, 20 µm.
    Flag Tagged Mouse Gapdh Cdna, supplied by OriGene, used in various techniques. Bioz Stars score: 86/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    In vivo transfection of cerebellar cells with Gfi1 / c-MYC and Otx2 / c-MYC induces Group 3 MB. a DAPI staining and GFP immunofluorescence of CD1 mouse brain section 3 months after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. b DAPI and GFP immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. c Hematoxylin and Flag (Gfi1) immunohistochemistry of CD1 mouse tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. d DAPI and Otx2 immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. e , f Hematoxylin and NPR3 immunohistochemistry of CD1 mice tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( e ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( f ). g , h GFP and GFAP immunofluorescence of tumors in CD1 mice after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( g ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( h ). i In vivo bioluminescent imaging of the upper body of a CD1 mouse transfected with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0, with a plasmid encoding firefly luciferase (pPBLuc). Imaging was performed weekly from the sixth week after transfection. j In vivo bioluminescent imaging of the lumbosacral region of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. The luminescence signal is expressed in photons/sec/mm2. k DAPI and immunofluorescence for pH3 of transversal section of the sacral portion of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l DAPI and immunofluorescence for GFP and pH3 of sacral transversal section of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l is the higher magnification of the white square in ( k ). Scale bars 1 mm in ( a , b ), 100 µm in ( c – h ), 500 µm in ( k ).

    Journal: Nature Communications

    Article Title: Modeling medulloblastoma in vivo and with human cerebellar organoids

    doi: 10.1038/s41467-019-13989-3

    Figure Lengend Snippet: In vivo transfection of cerebellar cells with Gfi1 / c-MYC and Otx2 / c-MYC induces Group 3 MB. a DAPI staining and GFP immunofluorescence of CD1 mouse brain section 3 months after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. b DAPI and GFP immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. c Hematoxylin and Flag (Gfi1) immunohistochemistry of CD1 mouse tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0. d DAPI and Otx2 immunofluorescence of CD1 mouse brain section 1 month after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0. e , f Hematoxylin and NPR3 immunohistochemistry of CD1 mice tumors after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( e ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( f ). g , h GFP and GFAP immunofluorescence of tumors in CD1 mice after transfection with pPBase + pPBMyc + pPBGfi1 + pPBVenus at P0 ( g ) and with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0 ( h ). i In vivo bioluminescent imaging of the upper body of a CD1 mouse transfected with pPBase + pPBMyc + pPBOtx2 + pPBVenus at P0, with a plasmid encoding firefly luciferase (pPBLuc). Imaging was performed weekly from the sixth week after transfection. j In vivo bioluminescent imaging of the lumbosacral region of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. The luminescence signal is expressed in photons/sec/mm2. k DAPI and immunofluorescence for pH3 of transversal section of the sacral portion of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l DAPI and immunofluorescence for GFP and pH3 of sacral transversal section of the spine of a CD1 mouse 8 weeks after transfection with pPBase + pPBMyc + pPBOtx2 + pPBVenus + pPBLuc at P0. l is the higher magnification of the white square in ( k ). Scale bars 1 mm in ( a , b ), 100 µm in ( c – h ), 500 µm in ( k ).

    Article Snippet: FLAG-tagged Gfi1 cDNA (Addgene #44630) was subcloned into the piggyBac donor backbone together with a IRES-GFP cassette, generating the plasmid pPB CAG Gfi1:FLAG-IRES-GFP.

    Techniques: In Vivo, Transfection, Staining, Immunofluorescence, Immunohistochemistry, Mouse Assay, Imaging, Plasmid Preparation, Luciferase

    Genetic experiments establish an essential LKB1-SIK3-HDAC4 pathway in AML (A) SIK3 mutants used in this study. (B) Western blot in the cells transduced with empty vector or the indicated SIK3 cDNA. (C and D) Competition-based proliferation assays in the cells transduced with empty vector or the indicated SIK3 cDNA and infected with the indicated GFP-linked sgRNA.. (E) Chemical structure of HG-9-91-01. (F – H) Relative growth of the indicated cells, harboring empty vector, SIK3, or SIK3 T142Q cDNA, upon HG-9-91-01 treatment. Normalized relative luminescence unit (RLU) was shown after 3 days culture with DMSO (0.1%) or HG-9-91-01 at the indicated concentrations. The mean ± SEM (n=3) and four-parameter dose-response curves are plotted. (I) Competition-based proliferation assays in the cells co-infected with GFP-linked sgRNA and mCherry-linked sgRNA. The percentage of double mCherry+/GFP+ cells is shown. (J) Western blot in the cells infected with the indicated sgRNA. (K) Western blot in the cells harboring empty vector, SIK3, or SIK3 T142Q cDNA, followed by treatment with DMSO (0.1%) or 100 nM HG-9-91-01 for 2 hrs. All bar graphs represent the mean ± SEM (n=3). Neg1, Neg2: negative controls. .

    Journal: Molecular cell

    Article Title: LKB1, Salt-Inducible Kinases, and MEF2C are linked dependencies in acute myeloid leukemia

    doi: 10.1016/j.molcel.2018.02.011

    Figure Lengend Snippet: Genetic experiments establish an essential LKB1-SIK3-HDAC4 pathway in AML (A) SIK3 mutants used in this study. (B) Western blot in the cells transduced with empty vector or the indicated SIK3 cDNA. (C and D) Competition-based proliferation assays in the cells transduced with empty vector or the indicated SIK3 cDNA and infected with the indicated GFP-linked sgRNA.. (E) Chemical structure of HG-9-91-01. (F – H) Relative growth of the indicated cells, harboring empty vector, SIK3, or SIK3 T142Q cDNA, upon HG-9-91-01 treatment. Normalized relative luminescence unit (RLU) was shown after 3 days culture with DMSO (0.1%) or HG-9-91-01 at the indicated concentrations. The mean ± SEM (n=3) and four-parameter dose-response curves are plotted. (I) Competition-based proliferation assays in the cells co-infected with GFP-linked sgRNA and mCherry-linked sgRNA. The percentage of double mCherry+/GFP+ cells is shown. (J) Western blot in the cells infected with the indicated sgRNA. (K) Western blot in the cells harboring empty vector, SIK3, or SIK3 T142Q cDNA, followed by treatment with DMSO (0.1%) or 100 nM HG-9-91-01 for 2 hrs. All bar graphs represent the mean ± SEM (n=3). Neg1, Neg2: negative controls. .

    Article Snippet: Flag-tagged LKB1 cDNA (addgene: #8590) ( ) was cloned into LentiV_Neo vector.

    Techniques: Western Blot, Transduction, Plasmid Preparation, Infection

    IDO impairs antitumor effects of anti-CTLA4 in the context of B16 tumors engineered to overexpress IDO. (A) Expression of GFP (black line) or IDO/GFP (gray) in B16F10 cells determined by flow cytometry. WT B16F10 cells were used for comparison (dotted line). (B) In vitro growth rate of GFP-B16F10 and IDO/GFP-B16F10 cells. (C) Mean growth of GFP-B16F10 and IDO/GFP-B16F10 tumors in naive and irradiated (450 cGy) mice. (D) Tumor-free survival curves for C57BL/6 mice challenged with GFP-B16F10 or IDO/GFP-B16F10 tumor cells i.d. and treated with anti–CTLA-4 or anti-CLTA-4/1MT. Data represent cumulative results from two independent experiments with five mice/group. Statistical significance was determined with Log-Rank test (*, P

    Journal: The Journal of Experimental Medicine

    Article Title: Indoleamine 2,3-dioxygenase is a critical resistance mechanism in antitumor T cell immunotherapy targeting CTLA-4

    doi: 10.1084/jem.20130066

    Figure Lengend Snippet: IDO impairs antitumor effects of anti-CTLA4 in the context of B16 tumors engineered to overexpress IDO. (A) Expression of GFP (black line) or IDO/GFP (gray) in B16F10 cells determined by flow cytometry. WT B16F10 cells were used for comparison (dotted line). (B) In vitro growth rate of GFP-B16F10 and IDO/GFP-B16F10 cells. (C) Mean growth of GFP-B16F10 and IDO/GFP-B16F10 tumors in naive and irradiated (450 cGy) mice. (D) Tumor-free survival curves for C57BL/6 mice challenged with GFP-B16F10 or IDO/GFP-B16F10 tumor cells i.d. and treated with anti–CTLA-4 or anti-CLTA-4/1MT. Data represent cumulative results from two independent experiments with five mice/group. Statistical significance was determined with Log-Rank test (*, P

    Article Snippet: GFP-tagged murine IDO cDNA (OriGene Technologies) was cloned into the pMDG lentiviral vector.

    Techniques: Expressing, Flow Cytometry, Cytometry, In Vitro, Irradiation, Mouse Assay

    Characterization of the GFAP c.1289G > A/p.R430H mutation. A : Schematic representation of the exonic structure of different GFAP isoforms. Dotted lines indicate the termination codons. The arrows indicate the position of the c.1289G > A variant (Note that in GFAP-κ the c.1289G > A mutation is part of the 3′-UTR). B : Electropherograms of GFAP exon 7A region containing c.1289G > A variant, in patients 1 and 2 (Pt1, Pt2) and in their mother (I-2). C : The histogram displays the percentages of cells transfected with GFP-GFAP-ϵ wt (green bars) or GFP-GFAP-ϵ R430H (purple bars), classified in filamentous pattern (F), cytoplasmic aggregates on a filamentous pattern (F + A), cytoplasmic aggregates with no filamentous pattern (A). Scale bars represent 15 μm. A total of 324 cells for GFP-GFAP-ϵ wt and 285 for GFP-GFAP-ϵ R430H , from 3 independent experiments, were blindly analyzed by two different operators. ANOVA test for interaction p = 0.001.

    Journal: Orphanet Journal of Rare Diseases

    Article Title: Adult-onset Alexander disease, associated with a mutation in an alternative GFAP transcript, may be phenotypically modulated by a non-neutral HDAC6 variant

    doi: 10.1186/1750-1172-8-66

    Figure Lengend Snippet: Characterization of the GFAP c.1289G > A/p.R430H mutation. A : Schematic representation of the exonic structure of different GFAP isoforms. Dotted lines indicate the termination codons. The arrows indicate the position of the c.1289G > A variant (Note that in GFAP-κ the c.1289G > A mutation is part of the 3′-UTR). B : Electropherograms of GFAP exon 7A region containing c.1289G > A variant, in patients 1 and 2 (Pt1, Pt2) and in their mother (I-2). C : The histogram displays the percentages of cells transfected with GFP-GFAP-ϵ wt (green bars) or GFP-GFAP-ϵ R430H (purple bars), classified in filamentous pattern (F), cytoplasmic aggregates on a filamentous pattern (F + A), cytoplasmic aggregates with no filamentous pattern (A). Scale bars represent 15 μm. A total of 324 cells for GFP-GFAP-ϵ wt and 285 for GFP-GFAP-ϵ R430H , from 3 independent experiments, were blindly analyzed by two different operators. ANOVA test for interaction p = 0.001.

    Article Snippet: A GFP tagged GFAP cDNA (Origene RG225707) was modified by using Quick-change Site-directed mutagenesis kit (Stratagene) to introduce either the c.1289G > A or the c.1288C > T nucleotide change in the RG225707 clone, using primers listed in Additional file .

    Techniques: Mutagenesis, Variant Assay, Transfection

    uPARAP forms a complex with VEGFR-2 and VEGFR-3 and restricts VEGFR-2/VEGFR-3 heterodimerisation. In situ PLA in LECs ( a , b , e – i ) or PAECs ( d ) stimulated or not for 5 (T5) and 30 min (T30) with VEGF-C ( a , b , e – g ). a , b Interaction between uPARAP and VEGFR-2 ( a ) or VEGFR-3 ( b ). Results are those of one representative assay out of 3. Data represent mean (biological duplicates, 20 images analysed for quantification) ± SEM. Bars = 10 µm. c Immunoprecipitation (IP) of uPARAP at the indicated time point of VEGF-C stimulation. Western blots were revealed with an antibody (IB) against VEGFR-2, VEGFR-3 (upper panel) or uPARAP (lower panel). GAPDH was used as loading control. d In situ PLA detection of uPARAP interaction with VEGFR-2 or VEGFR-3 under basal conditions, in PAECs expressing VEGFR-2 (PAEC-VEGFR-2), VEGFR-3 (PAEC-VEGFR-3) or no VEGFR (PAEC). Bars = 10 µm. e , f In situ PLA detection of VEGFR-2/VEGFR-3 heterodimerisation in control LECs (Ctr), LECs deficient for uPARAP expression (siU1 and siU2) ( e ), or LECs overexpressing uPARAP (uPARAP cDNA) ( f ). g VEGFR-3 homodimerisation in LECs transfected with a VEGFR-3-Flag-tagged cDNA. Data correspond to the mean (biological triplicates, 60 images analysed for quantification, > 100 cells analysed per condition) ± SEM ( e – g ). h In situ PLA detection of uPARAP/VEGFR-2 and uPARAP/VEGFR-3 complexes in LECs migrating in a VEGF-C gradient. Bars = 20 µm. i In situ PLA detection of VEGFR-2/VEGFR-3 heterodimers in control LECs (Ctr) and uPARAP silenced LECs (siU) migrating in a VEGF-C gradient. The left histogram corresponds to the mean number of dots per cell. The right histogram represents PLA signal localisation with respect to the nucleus (one representative experiment out of 3 with 25 cells analysed per condition in each experiment). Bars = 20 µm. Statistical analyses were performed using a non-parametric Mann–Whitney test. i Chi-square test was used. * P

    Journal: Nature Communications

    Article Title: uPARAP/Endo180 receptor is a gatekeeper of VEGFR-2/VEGFR-3 heterodimerisation during pathological lymphangiogenesis

    doi: 10.1038/s41467-018-07514-1

    Figure Lengend Snippet: uPARAP forms a complex with VEGFR-2 and VEGFR-3 and restricts VEGFR-2/VEGFR-3 heterodimerisation. In situ PLA in LECs ( a , b , e – i ) or PAECs ( d ) stimulated or not for 5 (T5) and 30 min (T30) with VEGF-C ( a , b , e – g ). a , b Interaction between uPARAP and VEGFR-2 ( a ) or VEGFR-3 ( b ). Results are those of one representative assay out of 3. Data represent mean (biological duplicates, 20 images analysed for quantification) ± SEM. Bars = 10 µm. c Immunoprecipitation (IP) of uPARAP at the indicated time point of VEGF-C stimulation. Western blots were revealed with an antibody (IB) against VEGFR-2, VEGFR-3 (upper panel) or uPARAP (lower panel). GAPDH was used as loading control. d In situ PLA detection of uPARAP interaction with VEGFR-2 or VEGFR-3 under basal conditions, in PAECs expressing VEGFR-2 (PAEC-VEGFR-2), VEGFR-3 (PAEC-VEGFR-3) or no VEGFR (PAEC). Bars = 10 µm. e , f In situ PLA detection of VEGFR-2/VEGFR-3 heterodimerisation in control LECs (Ctr), LECs deficient for uPARAP expression (siU1 and siU2) ( e ), or LECs overexpressing uPARAP (uPARAP cDNA) ( f ). g VEGFR-3 homodimerisation in LECs transfected with a VEGFR-3-Flag-tagged cDNA. Data correspond to the mean (biological triplicates, 60 images analysed for quantification, > 100 cells analysed per condition) ± SEM ( e – g ). h In situ PLA detection of uPARAP/VEGFR-2 and uPARAP/VEGFR-3 complexes in LECs migrating in a VEGF-C gradient. Bars = 20 µm. i In situ PLA detection of VEGFR-2/VEGFR-3 heterodimers in control LECs (Ctr) and uPARAP silenced LECs (siU) migrating in a VEGF-C gradient. The left histogram corresponds to the mean number of dots per cell. The right histogram represents PLA signal localisation with respect to the nucleus (one representative experiment out of 3 with 25 cells analysed per condition in each experiment). Bars = 20 µm. Statistical analyses were performed using a non-parametric Mann–Whitney test. i Chi-square test was used. * P

    Article Snippet: LECs were transfected with uPARAP cDNA for uPARAP overexpression assays and with VEGFR-3 Flag-tagged cDNA (HG10806-NF, Sino Biological) for homodimers PLA assays using Viromer Yellow (VY-01LB, Lipocalyx) as transfection reagent.

    Techniques: In Situ, Proximity Ligation Assay, Immunoprecipitation, Western Blot, Expressing, Transfection, MANN-WHITNEY

    uPARAP downregulation impairs the Crk/JNK/paxillin pathway. a , b , d – f LECs were treated with siRNAs targeting uPARAP (siU1 and siU2) or a control siRNA (Ctr) and stimulated for the indicated time (minutes) with VEGF-C. a – f Western blot of total and phosphorylated (p) VEGFR-2 (Y1175), VEGFR-3 (Y1230/31) ( a ), ERK, AKT, JNK ( b , c ), Crk ( d, e ) and Paxillin (Y118, S178) ( f ). c PAECs expressing uPARAP were transfected with a plasmid carrying an intact (WT) or mutated (VEGFR-3 Y1063/F ) VEGFR-3 cDNA. JNK phosphorylation was evaluated by Western blot after VEGF-C stimulation. d , e LECs were treated (+ZM) or not (-ZM) with VEGFR-2 inhibitor (ZM323881), or siRNA against VEGFR-2 (siR2). For quantification of Western blot band densities, values were calculated as the ratio of phosphorylated to total form and normalised to the values obtained with the control (Ctr) ( n = 3, biological replicates). Statistical analyses were performed using a non-parametric Mann–Whitney test. * P

    Journal: Nature Communications

    Article Title: uPARAP/Endo180 receptor is a gatekeeper of VEGFR-2/VEGFR-3 heterodimerisation during pathological lymphangiogenesis

    doi: 10.1038/s41467-018-07514-1

    Figure Lengend Snippet: uPARAP downregulation impairs the Crk/JNK/paxillin pathway. a , b , d – f LECs were treated with siRNAs targeting uPARAP (siU1 and siU2) or a control siRNA (Ctr) and stimulated for the indicated time (minutes) with VEGF-C. a – f Western blot of total and phosphorylated (p) VEGFR-2 (Y1175), VEGFR-3 (Y1230/31) ( a ), ERK, AKT, JNK ( b , c ), Crk ( d, e ) and Paxillin (Y118, S178) ( f ). c PAECs expressing uPARAP were transfected with a plasmid carrying an intact (WT) or mutated (VEGFR-3 Y1063/F ) VEGFR-3 cDNA. JNK phosphorylation was evaluated by Western blot after VEGF-C stimulation. d , e LECs were treated (+ZM) or not (-ZM) with VEGFR-2 inhibitor (ZM323881), or siRNA against VEGFR-2 (siR2). For quantification of Western blot band densities, values were calculated as the ratio of phosphorylated to total form and normalised to the values obtained with the control (Ctr) ( n = 3, biological replicates). Statistical analyses were performed using a non-parametric Mann–Whitney test. * P

    Article Snippet: LECs were transfected with uPARAP cDNA for uPARAP overexpression assays and with VEGFR-3 Flag-tagged cDNA (HG10806-NF, Sino Biological) for homodimers PLA assays using Viromer Yellow (VY-01LB, Lipocalyx) as transfection reagent.

    Techniques: Western Blot, Expressing, Transfection, Plasmid Preparation, MANN-WHITNEY

    PTEN/NLK synthetic lethality is abrogated by FOXO1 silencing. ( A ) Survival analysis of HCT116 cells transfected with NLK and/or FOXO1 siRNA. HCT116 PTEN −/− and HCT116 PTEN +/+ cells were transfected with siRNA targeting NLK and FOXO1 as shown and surviving fractions determined after five days. The p value (*) was calculated using Student's t test. ( B ) Nuclear localisation of FOXO1 is enhanced in PTEN deficient cells upon NLK silencing. HCT116 isogenic cells were co-transfected with GFP-tagged FOXO1 cDNA in addition to control (non targeting) siRNA (siCON) or NLK siRNA. Two days later cells were fixed and stained with DAPI. Green signal represents GFP-FOXO1 and blue signal represents nuclear DAPI (nuclear) staining. Arrows indicate cells with nuclear localisation of FOXO1. ( C ) Senescence is increased by NLK siRNA in PTEN deficient cells. Bar chart of relative relative senescence levels caused by NLK silencing are shown. HCT116-derived PTEN isogenic cell lines were reversed transfected with a pool of two validated siRNAs against NLK, as well as siCON pool#2 (Dharmacon) as non-targeting control, using RNAiMAX (Invitrogen). Seven days after transfection cells were fixed and incubated overnight at 37°C in a solution containing X-gal. ( D ) Representative images for β-Galactosidase staining of PTEN deficient cells. Blue staining indicates β-Galactosidase.

    Journal: PLoS ONE

    Article Title: NLK Is a Novel Therapeutic Target for PTEN Deficient Tumour Cells

    doi: 10.1371/journal.pone.0047249

    Figure Lengend Snippet: PTEN/NLK synthetic lethality is abrogated by FOXO1 silencing. ( A ) Survival analysis of HCT116 cells transfected with NLK and/or FOXO1 siRNA. HCT116 PTEN −/− and HCT116 PTEN +/+ cells were transfected with siRNA targeting NLK and FOXO1 as shown and surviving fractions determined after five days. The p value (*) was calculated using Student's t test. ( B ) Nuclear localisation of FOXO1 is enhanced in PTEN deficient cells upon NLK silencing. HCT116 isogenic cells were co-transfected with GFP-tagged FOXO1 cDNA in addition to control (non targeting) siRNA (siCON) or NLK siRNA. Two days later cells were fixed and stained with DAPI. Green signal represents GFP-FOXO1 and blue signal represents nuclear DAPI (nuclear) staining. Arrows indicate cells with nuclear localisation of FOXO1. ( C ) Senescence is increased by NLK siRNA in PTEN deficient cells. Bar chart of relative relative senescence levels caused by NLK silencing are shown. HCT116-derived PTEN isogenic cell lines were reversed transfected with a pool of two validated siRNAs against NLK, as well as siCON pool#2 (Dharmacon) as non-targeting control, using RNAiMAX (Invitrogen). Seven days after transfection cells were fixed and incubated overnight at 37°C in a solution containing X-gal. ( D ) Representative images for β-Galactosidase staining of PTEN deficient cells. Blue staining indicates β-Galactosidase.

    Article Snippet: FOXO1 detection HCT116 isogenic cells were co-transfected with GFP-tagged FOXO1 cDNA (Origene) in addition to control (non targeting) siRNA (siCON) or NLK siRNA.

    Techniques: Transfection, Staining, Derivative Assay, Incubation

    Primer integrity and specificity analysis. ( A ) 2% agarose gel electrophoresis of control 8093 RT/PCR products using gapdh , 18 s rRNA , and cassini shows a single band for each target gene. ( B ) Real time RT/PCR on RNA isolated from non-transfected MEFs (control), MEFs transfected with an EGFP- cassini construct or with empty EGFP-C1 plasmid. Values (mean ± SD of triplicate real-time PCR) are expressed as percent of gapdh . ( C, D ) Derivative dissociation curves from real-time RT/PCR on mouse ( C ) and human ( D ) RNA confirm single product amplification. The cassini amplicon shows a single peak for both murine and human RNA with a T m of ~76°C. Gapdh primers peak at T m ~85.5°C. ***p

    Journal: BMC Genomics

    Article Title: Expression of cassini, a murine gamma-satellite sequence conserved in evolution, is regulated in normal and malignant hematopoietic cells

    doi: 10.1186/1471-2164-13-418

    Figure Lengend Snippet: Primer integrity and specificity analysis. ( A ) 2% agarose gel electrophoresis of control 8093 RT/PCR products using gapdh , 18 s rRNA , and cassini shows a single band for each target gene. ( B ) Real time RT/PCR on RNA isolated from non-transfected MEFs (control), MEFs transfected with an EGFP- cassini construct or with empty EGFP-C1 plasmid. Values (mean ± SD of triplicate real-time PCR) are expressed as percent of gapdh . ( C, D ) Derivative dissociation curves from real-time RT/PCR on mouse ( C ) and human ( D ) RNA confirm single product amplification. The cassini amplicon shows a single peak for both murine and human RNA with a T m of ~76°C. Gapdh primers peak at T m ~85.5°C. ***p

    Article Snippet: Protein product of EGFP-tagged cassini cDNA.

    Techniques: Agarose Gel Electrophoresis, Reverse Transcription Polymerase Chain Reaction, Quantitative RT-PCR, Isolation, Transfection, Construct, Plasmid Preparation, Real-time Polymerase Chain Reaction, Amplification

    Increased intracellular sodium inhibits HDAC6 activity. a DAOY cells were incubated with increasing concentrations of ouabain for three hours and then lysed for an in vitro HDAC activity assay as described in Materials and Methods. b DAOY cells were incubated with indicated concentrations of ouabain and the protein levels of the indicated HDAC isoforms were determined by immunoblotting. Actin immunoblot ensures equal loading. c After incubation with indicated concentrations of gramicidin A, HDAC6 was immunoprecipitated with a specific antibody and then subjected to an in vitro HDAC activity assay using a fluorescent substrate. d DAOY cells were transiently transfected with Flag-HDAC6 cDNA. The cells were incubated with the indicated concentrations of gramicidin A followed by immunoprecipitation of HDAC6. The immunoprecipitates were washed extensively and then incubated with polymerized microtubules to measure deacetylase activity. The reaction was terminated by adding lysis buffer and the samples were processed for immunoblotting with indicated antibodies. e Immunoblot for HDAC6 of stable clones of DAOY cells expressing HDAC6 shRNA. Control cells were transfected with vector only and processed in parallel. A GAPDH blot ensures that equal amounts of protein were used for analysis. f The HDAC6 knockdown cells were incubated with either High Na + or Low Na + buffer and the trafficking patterns of EGF-positive vesicles were analyzed as described in figure legend 3. *, P

    Journal: BMC Cell Biology

    Article Title: EGF-induced sodium influx regulates EGFR trafficking through HDAC6 and tubulin acetylation

    doi: 10.1186/s12860-015-0070-8

    Figure Lengend Snippet: Increased intracellular sodium inhibits HDAC6 activity. a DAOY cells were incubated with increasing concentrations of ouabain for three hours and then lysed for an in vitro HDAC activity assay as described in Materials and Methods. b DAOY cells were incubated with indicated concentrations of ouabain and the protein levels of the indicated HDAC isoforms were determined by immunoblotting. Actin immunoblot ensures equal loading. c After incubation with indicated concentrations of gramicidin A, HDAC6 was immunoprecipitated with a specific antibody and then subjected to an in vitro HDAC activity assay using a fluorescent substrate. d DAOY cells were transiently transfected with Flag-HDAC6 cDNA. The cells were incubated with the indicated concentrations of gramicidin A followed by immunoprecipitation of HDAC6. The immunoprecipitates were washed extensively and then incubated with polymerized microtubules to measure deacetylase activity. The reaction was terminated by adding lysis buffer and the samples were processed for immunoblotting with indicated antibodies. e Immunoblot for HDAC6 of stable clones of DAOY cells expressing HDAC6 shRNA. Control cells were transfected with vector only and processed in parallel. A GAPDH blot ensures that equal amounts of protein were used for analysis. f The HDAC6 knockdown cells were incubated with either High Na + or Low Na + buffer and the trafficking patterns of EGF-positive vesicles were analyzed as described in figure legend 3. *, P

    Article Snippet: Flag-tagged HDAC6 cDNA was obtained from Addgene (Cambridge, MA) and transfected into human medulloblastoma DAOY cells (ATCC, Rockville, MD) using Lipofectamine 2000 (Invitrogen).

    Techniques: Activity Assay, Incubation, In Vitro, HDAC Activity Assay, Immunoprecipitation, Transfection, Histone Deacetylase Assay, Lysis, Clone Assay, Expressing, shRNA, Plasmid Preparation

    DDC-induced ROS formation is associated with a depletion of energy metabolism and antioxidant enzymes and is regulated by GAPDH. (A) Hepatocytes from C3H and C57BL mice were left untreated (−) or treated with either 0.1% DMSO vehicle (0) or the indicated concentrations of DDC for 48 h. Equal protein amounts (NP-40 lysates) were analyzed for expression of the indicated proteins. Samples from each strain were analyzed on separate gels. PDIA4 levels were unaltered (loading control). (B) C57BL hepatocytes were transfected with Control or GAPDH siRNA for 24 h and were then cultured for an additional 24 h in the presence of 0.025% DMSO vehicle (0) or 100 µM DDC or were left untreated (−). The NP-40 cell lysates were analyzed for the expression of the various proteins indicated. (C) Overexpression of Flag-tagged mouse GAPDH in isolated hepatocytes. NP-40 lysates were analyzed for expression of Flag-tagged GAPDH and total GAPDH. The Coomassie stain is included as a loading control. (D) Flag-GAPDH expression in C57BL hepatocytes, as determined by immunostaining with a mouse anti-Flag antibody (representing overexpressed GAPDH; green) and a rabbit anti-GAPDH antibody (representing total GAPDH; red). (E) C57BL hepatocytes were mock transfected (Control) or transfected with GAPDH siRNA or Flag-GAPDH mouse cDNA for 24 h and were then treated with 100 µM DDC for an additional 24 h. Representative images of ROS signal (green) with DAPI nuclear counterstain (blue) are shown. ROS levels (quantified as described in Materials and methods) exhibited a 2.2-fold increase after GAPDH knockdown and a 10-fold decrease after GAPDH overexpression relative to control. Bars, 20 µm.

    Journal: The Journal of Cell Biology

    Article Title: Energy determinants GAPDH and NDPK act as genetic modifiers for hepatocyte inclusion formation

    doi: 10.1083/jcb.201102142

    Figure Lengend Snippet: DDC-induced ROS formation is associated with a depletion of energy metabolism and antioxidant enzymes and is regulated by GAPDH. (A) Hepatocytes from C3H and C57BL mice were left untreated (−) or treated with either 0.1% DMSO vehicle (0) or the indicated concentrations of DDC for 48 h. Equal protein amounts (NP-40 lysates) were analyzed for expression of the indicated proteins. Samples from each strain were analyzed on separate gels. PDIA4 levels were unaltered (loading control). (B) C57BL hepatocytes were transfected with Control or GAPDH siRNA for 24 h and were then cultured for an additional 24 h in the presence of 0.025% DMSO vehicle (0) or 100 µM DDC or were left untreated (−). The NP-40 cell lysates were analyzed for the expression of the various proteins indicated. (C) Overexpression of Flag-tagged mouse GAPDH in isolated hepatocytes. NP-40 lysates were analyzed for expression of Flag-tagged GAPDH and total GAPDH. The Coomassie stain is included as a loading control. (D) Flag-GAPDH expression in C57BL hepatocytes, as determined by immunostaining with a mouse anti-Flag antibody (representing overexpressed GAPDH; green) and a rabbit anti-GAPDH antibody (representing total GAPDH; red). (E) C57BL hepatocytes were mock transfected (Control) or transfected with GAPDH siRNA or Flag-GAPDH mouse cDNA for 24 h and were then treated with 100 µM DDC for an additional 24 h. Representative images of ROS signal (green) with DAPI nuclear counterstain (blue) are shown. ROS levels (quantified as described in Materials and methods) exhibited a 2.2-fold increase after GAPDH knockdown and a 10-fold decrease after GAPDH overexpression relative to control. Bars, 20 µm.

    Article Snippet: Transfection of Flag-tagged mouse GAPDH cDNA (OriGene) was performed using Lipofectamine LTX (Invitrogen) according to the manufacturer’s instructions. siRNA and DNA-transfected hepatocytes were exposed to 100 µM DDC for 24 h.

    Techniques: Mouse Assay, Expressing, Transfection, Cell Culture, Over Expression, Isolation, Staining, Immunostaining