bamhi Origene Search Results


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  • 95
    New England Biolabs restriction enzymes bamhi
    Restriction Enzymes Bamhi, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 95/100, based on 332 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 95 stars, based on 332 article reviews
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    restriction enzymes bamhi - by Bioz Stars, 2020-02
    95/100 stars
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    95
    Millipore bamh
    Bamh, supplied by Millipore, used in various techniques. Bioz Stars score: 95/100, based on 441 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 95 stars, based on 441 article reviews
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    bamh - by Bioz Stars, 2020-02
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    90
    Thermo Fisher bamhi
    Direct visualization of <t>HPV88</t> on an EtBr-stained gel, after digestion with <t>BamHI,</t> lane 1. Lane M, Gene Ruler 1 kb DNA ladder (Fermentas).
    Bamhi, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 10438 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 90 stars, based on 10438 article reviews
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    79
    OriGene bamhi hindiii cloning sites
    Direct visualization of <t>HPV88</t> on an EtBr-stained gel, after digestion with <t>BamHI,</t> lane 1. Lane M, Gene Ruler 1 kb DNA ladder (Fermentas).
    Bamhi Hindiii Cloning Sites, supplied by OriGene, used in various techniques. Bioz Stars score: 79/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    83
    OriGene pet15b hsspata22 1 363
    Direct visualization of <t>HPV88</t> on an EtBr-stained gel, after digestion with <t>BamHI,</t> lane 1. Lane M, Gene Ruler 1 kb DNA ladder (Fermentas).
    Pet15b Hsspata22 1 363, supplied by OriGene, used in various techniques. Bioz Stars score: 83/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    pet15b hsspata22 1 363 - by Bioz Stars, 2020-02
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    90
    Thermo Fisher bamh i
    Direct visualization of <t>HPV88</t> on an EtBr-stained gel, after digestion with <t>BamHI,</t> lane 1. Lane M, Gene Ruler 1 kb DNA ladder (Fermentas).
    Bamh I, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 1161 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    bamh i - by Bioz Stars, 2020-02
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    96
    Thermo Fisher ecori
    Direct visualization of <t>HPV88</t> on an EtBr-stained gel, after digestion with <t>BamHI,</t> lane 1. Lane M, Gene Ruler 1 kb DNA ladder (Fermentas).
    Ecori, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 96/100, based on 9775 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 96 stars, based on 9775 article reviews
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    89
    OriGene c terminal myc
    Direct visualization of <t>HPV88</t> on an EtBr-stained gel, after digestion with <t>BamHI,</t> lane 1. Lane M, Gene Ruler 1 kb DNA ladder (Fermentas).
    C Terminal Myc, supplied by OriGene, used in various techniques. Bioz Stars score: 89/100, based on 84 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    79
    OriGene cdna source
    Direct visualization of <t>HPV88</t> on an EtBr-stained gel, after digestion with <t>BamHI,</t> lane 1. Lane M, Gene Ruler 1 kb DNA ladder (Fermentas).
    Cdna Source, supplied by OriGene, used in various techniques. Bioz Stars score: 79/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    78
    OriGene cdc25a plasmid
    FOXM1 directly binds to the putative FOXM1 consensus binding sequences located on the <t>CDC25A</t> promoter. (A) FOXM1 binding to CDC25A promoter was confirmed by ChIP-qPCR. Primer set A was designed to span three FOXM1 binding sites, and primer set B, targeting a region of the promoter that did not include the FOXM1 binding sites, was used as the control. Chromatin immunoprecipitations of U2OS cells were prepared using anti-FOXM1 antibody. Semi-quantitative PCR was performed using the primer sets A and B, and the PCR products were detected by electrophoresis (Top). Quantitative PCR was performed using the primer sets A and B, and the binding activity of FOXM1 to CDC25A promoter was evaluated by the fold enrichment method. (B–C) The three FOXM1 binding sites on the CDC25A promoter are functionally redundant (B) Schematic diagram showing the mutations of the three FOXM1 binding sites on the CDC25A promoter. (C) U2OS cells were transfected with pACT-FOXM1 or empty vector control, along with the indicated CDC25A promoter-luciferase reporter construct. Targeted mutation of single FOXM1 binding sites did not significantly affect CDC25A transactivation. Mutations of sites 1 and 2, sites 2 and 3, and sites 1, 2, and 3 significantly reduced the transactivation of the CDC25A-luciferase reporter. Data were normalized to Renilla luciferase activities and are presented as the mean wild-typefold induction over empty vector control ± SD (N = 3). Data were subjected to one-way ANOVA (significance level α = 0.05) and Dunnett's multi-comparison post-hoc tests (mutations vs. wild-type). * P
    Cdc25a Plasmid, supplied by OriGene, used in various techniques. Bioz Stars score: 78/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 78 stars, based on 8 article reviews
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    81
    OriGene human peroxisomal membrane protein
    FOXM1 directly binds to the putative FOXM1 consensus binding sequences located on the <t>CDC25A</t> promoter. (A) FOXM1 binding to CDC25A promoter was confirmed by ChIP-qPCR. Primer set A was designed to span three FOXM1 binding sites, and primer set B, targeting a region of the promoter that did not include the FOXM1 binding sites, was used as the control. Chromatin immunoprecipitations of U2OS cells were prepared using anti-FOXM1 antibody. Semi-quantitative PCR was performed using the primer sets A and B, and the PCR products were detected by electrophoresis (Top). Quantitative PCR was performed using the primer sets A and B, and the binding activity of FOXM1 to CDC25A promoter was evaluated by the fold enrichment method. (B–C) The three FOXM1 binding sites on the CDC25A promoter are functionally redundant (B) Schematic diagram showing the mutations of the three FOXM1 binding sites on the CDC25A promoter. (C) U2OS cells were transfected with pACT-FOXM1 or empty vector control, along with the indicated CDC25A promoter-luciferase reporter construct. Targeted mutation of single FOXM1 binding sites did not significantly affect CDC25A transactivation. Mutations of sites 1 and 2, sites 2 and 3, and sites 1, 2, and 3 significantly reduced the transactivation of the CDC25A-luciferase reporter. Data were normalized to Renilla luciferase activities and are presented as the mean wild-typefold induction over empty vector control ± SD (N = 3). Data were subjected to one-way ANOVA (significance level α = 0.05) and Dunnett's multi-comparison post-hoc tests (mutations vs. wild-type). * P
    Human Peroxisomal Membrane Protein, supplied by OriGene, used in various techniques. Bioz Stars score: 81/100, based on 22 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    OriGene pcmv6
    FOXM1 directly binds to the putative FOXM1 consensus binding sequences located on the <t>CDC25A</t> promoter. (A) FOXM1 binding to CDC25A promoter was confirmed by ChIP-qPCR. Primer set A was designed to span three FOXM1 binding sites, and primer set B, targeting a region of the promoter that did not include the FOXM1 binding sites, was used as the control. Chromatin immunoprecipitations of U2OS cells were prepared using anti-FOXM1 antibody. Semi-quantitative PCR was performed using the primer sets A and B, and the PCR products were detected by electrophoresis (Top). Quantitative PCR was performed using the primer sets A and B, and the binding activity of FOXM1 to CDC25A promoter was evaluated by the fold enrichment method. (B–C) The three FOXM1 binding sites on the CDC25A promoter are functionally redundant (B) Schematic diagram showing the mutations of the three FOXM1 binding sites on the CDC25A promoter. (C) U2OS cells were transfected with pACT-FOXM1 or empty vector control, along with the indicated CDC25A promoter-luciferase reporter construct. Targeted mutation of single FOXM1 binding sites did not significantly affect CDC25A transactivation. Mutations of sites 1 and 2, sites 2 and 3, and sites 1, 2, and 3 significantly reduced the transactivation of the CDC25A-luciferase reporter. Data were normalized to Renilla luciferase activities and are presented as the mean wild-typefold induction over empty vector control ± SD (N = 3). Data were subjected to one-way ANOVA (significance level α = 0.05) and Dunnett's multi-comparison post-hoc tests (mutations vs. wild-type). * P
    Pcmv6, supplied by OriGene, used in various techniques. Bioz Stars score: 99/100, based on 380 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    80
    OriGene plasmid rc221178
    FOXM1 directly binds to the putative FOXM1 consensus binding sequences located on the <t>CDC25A</t> promoter. (A) FOXM1 binding to CDC25A promoter was confirmed by ChIP-qPCR. Primer set A was designed to span three FOXM1 binding sites, and primer set B, targeting a region of the promoter that did not include the FOXM1 binding sites, was used as the control. Chromatin immunoprecipitations of U2OS cells were prepared using anti-FOXM1 antibody. Semi-quantitative PCR was performed using the primer sets A and B, and the PCR products were detected by electrophoresis (Top). Quantitative PCR was performed using the primer sets A and B, and the binding activity of FOXM1 to CDC25A promoter was evaluated by the fold enrichment method. (B–C) The three FOXM1 binding sites on the CDC25A promoter are functionally redundant (B) Schematic diagram showing the mutations of the three FOXM1 binding sites on the CDC25A promoter. (C) U2OS cells were transfected with pACT-FOXM1 or empty vector control, along with the indicated CDC25A promoter-luciferase reporter construct. Targeted mutation of single FOXM1 binding sites did not significantly affect CDC25A transactivation. Mutations of sites 1 and 2, sites 2 and 3, and sites 1, 2, and 3 significantly reduced the transactivation of the CDC25A-luciferase reporter. Data were normalized to Renilla luciferase activities and are presented as the mean wild-typefold induction over empty vector control ± SD (N = 3). Data were subjected to one-way ANOVA (significance level α = 0.05) and Dunnett's multi-comparison post-hoc tests (mutations vs. wild-type). * P
    Plasmid Rc221178, supplied by OriGene, used in various techniques. Bioz Stars score: 80/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    OriGene pcmv6 entry vector
    FOXM1 directly binds to the putative FOXM1 consensus binding sequences located on the <t>CDC25A</t> promoter. (A) FOXM1 binding to CDC25A promoter was confirmed by ChIP-qPCR. Primer set A was designed to span three FOXM1 binding sites, and primer set B, targeting a region of the promoter that did not include the FOXM1 binding sites, was used as the control. Chromatin immunoprecipitations of U2OS cells were prepared using anti-FOXM1 antibody. Semi-quantitative PCR was performed using the primer sets A and B, and the PCR products were detected by electrophoresis (Top). Quantitative PCR was performed using the primer sets A and B, and the binding activity of FOXM1 to CDC25A promoter was evaluated by the fold enrichment method. (B–C) The three FOXM1 binding sites on the CDC25A promoter are functionally redundant (B) Schematic diagram showing the mutations of the three FOXM1 binding sites on the CDC25A promoter. (C) U2OS cells were transfected with pACT-FOXM1 or empty vector control, along with the indicated CDC25A promoter-luciferase reporter construct. Targeted mutation of single FOXM1 binding sites did not significantly affect CDC25A transactivation. Mutations of sites 1 and 2, sites 2 and 3, and sites 1, 2, and 3 significantly reduced the transactivation of the CDC25A-luciferase reporter. Data were normalized to Renilla luciferase activities and are presented as the mean wild-typefold induction over empty vector control ± SD (N = 3). Data were subjected to one-way ANOVA (significance level α = 0.05) and Dunnett's multi-comparison post-hoc tests (mutations vs. wild-type). * P
    Pcmv6 Entry Vector, supplied by OriGene, used in various techniques. Bioz Stars score: 90/100, based on 45 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    78
    OriGene pcas guide egfp
    FOXM1 directly binds to the putative FOXM1 consensus binding sequences located on the <t>CDC25A</t> promoter. (A) FOXM1 binding to CDC25A promoter was confirmed by ChIP-qPCR. Primer set A was designed to span three FOXM1 binding sites, and primer set B, targeting a region of the promoter that did not include the FOXM1 binding sites, was used as the control. Chromatin immunoprecipitations of U2OS cells were prepared using anti-FOXM1 antibody. Semi-quantitative PCR was performed using the primer sets A and B, and the PCR products were detected by electrophoresis (Top). Quantitative PCR was performed using the primer sets A and B, and the binding activity of FOXM1 to CDC25A promoter was evaluated by the fold enrichment method. (B–C) The three FOXM1 binding sites on the CDC25A promoter are functionally redundant (B) Schematic diagram showing the mutations of the three FOXM1 binding sites on the CDC25A promoter. (C) U2OS cells were transfected with pACT-FOXM1 or empty vector control, along with the indicated CDC25A promoter-luciferase reporter construct. Targeted mutation of single FOXM1 binding sites did not significantly affect CDC25A transactivation. Mutations of sites 1 and 2, sites 2 and 3, and sites 1, 2, and 3 significantly reduced the transactivation of the CDC25A-luciferase reporter. Data were normalized to Renilla luciferase activities and are presented as the mean wild-typefold induction over empty vector control ± SD (N = 3). Data were subjected to one-way ANOVA (significance level α = 0.05) and Dunnett's multi-comparison post-hoc tests (mutations vs. wild-type). * P
    Pcas Guide Egfp, supplied by OriGene, used in various techniques. Bioz Stars score: 78/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    78
    OriGene pcmv6 tpmt plasmid
    FOXM1 directly binds to the putative FOXM1 consensus binding sequences located on the <t>CDC25A</t> promoter. (A) FOXM1 binding to CDC25A promoter was confirmed by ChIP-qPCR. Primer set A was designed to span three FOXM1 binding sites, and primer set B, targeting a region of the promoter that did not include the FOXM1 binding sites, was used as the control. Chromatin immunoprecipitations of U2OS cells were prepared using anti-FOXM1 antibody. Semi-quantitative PCR was performed using the primer sets A and B, and the PCR products were detected by electrophoresis (Top). Quantitative PCR was performed using the primer sets A and B, and the binding activity of FOXM1 to CDC25A promoter was evaluated by the fold enrichment method. (B–C) The three FOXM1 binding sites on the CDC25A promoter are functionally redundant (B) Schematic diagram showing the mutations of the three FOXM1 binding sites on the CDC25A promoter. (C) U2OS cells were transfected with pACT-FOXM1 or empty vector control, along with the indicated CDC25A promoter-luciferase reporter construct. Targeted mutation of single FOXM1 binding sites did not significantly affect CDC25A transactivation. Mutations of sites 1 and 2, sites 2 and 3, and sites 1, 2, and 3 significantly reduced the transactivation of the CDC25A-luciferase reporter. Data were normalized to Renilla luciferase activities and are presented as the mean wild-typefold induction over empty vector control ± SD (N = 3). Data were subjected to one-way ANOVA (significance level α = 0.05) and Dunnett's multi-comparison post-hoc tests (mutations vs. wild-type). * P
    Pcmv6 Tpmt Plasmid, supplied by OriGene, used in various techniques. Bioz Stars score: 78/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    80
    OriGene pcmv6 ac rfp
    FOXM1 directly binds to the putative FOXM1 consensus binding sequences located on the <t>CDC25A</t> promoter. (A) FOXM1 binding to CDC25A promoter was confirmed by ChIP-qPCR. Primer set A was designed to span three FOXM1 binding sites, and primer set B, targeting a region of the promoter that did not include the FOXM1 binding sites, was used as the control. Chromatin immunoprecipitations of U2OS cells were prepared using anti-FOXM1 antibody. Semi-quantitative PCR was performed using the primer sets A and B, and the PCR products were detected by electrophoresis (Top). Quantitative PCR was performed using the primer sets A and B, and the binding activity of FOXM1 to CDC25A promoter was evaluated by the fold enrichment method. (B–C) The three FOXM1 binding sites on the CDC25A promoter are functionally redundant (B) Schematic diagram showing the mutations of the three FOXM1 binding sites on the CDC25A promoter. (C) U2OS cells were transfected with pACT-FOXM1 or empty vector control, along with the indicated CDC25A promoter-luciferase reporter construct. Targeted mutation of single FOXM1 binding sites did not significantly affect CDC25A transactivation. Mutations of sites 1 and 2, sites 2 and 3, and sites 1, 2, and 3 significantly reduced the transactivation of the CDC25A-luciferase reporter. Data were normalized to Renilla luciferase activities and are presented as the mean wild-typefold induction over empty vector control ± SD (N = 3). Data were subjected to one-way ANOVA (significance level α = 0.05) and Dunnett's multi-comparison post-hoc tests (mutations vs. wild-type). * P
    Pcmv6 Ac Rfp, supplied by OriGene, used in various techniques. Bioz Stars score: 80/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    79
    OriGene pgfp v rs cloning vector
    FOXM1 directly binds to the putative FOXM1 consensus binding sequences located on the <t>CDC25A</t> promoter. (A) FOXM1 binding to CDC25A promoter was confirmed by ChIP-qPCR. Primer set A was designed to span three FOXM1 binding sites, and primer set B, targeting a region of the promoter that did not include the FOXM1 binding sites, was used as the control. Chromatin immunoprecipitations of U2OS cells were prepared using anti-FOXM1 antibody. Semi-quantitative PCR was performed using the primer sets A and B, and the PCR products were detected by electrophoresis (Top). Quantitative PCR was performed using the primer sets A and B, and the binding activity of FOXM1 to CDC25A promoter was evaluated by the fold enrichment method. (B–C) The three FOXM1 binding sites on the CDC25A promoter are functionally redundant (B) Schematic diagram showing the mutations of the three FOXM1 binding sites on the CDC25A promoter. (C) U2OS cells were transfected with pACT-FOXM1 or empty vector control, along with the indicated CDC25A promoter-luciferase reporter construct. Targeted mutation of single FOXM1 binding sites did not significantly affect CDC25A transactivation. Mutations of sites 1 and 2, sites 2 and 3, and sites 1, 2, and 3 significantly reduced the transactivation of the CDC25A-luciferase reporter. Data were normalized to Renilla luciferase activities and are presented as the mean wild-typefold induction over empty vector control ± SD (N = 3). Data were subjected to one-way ANOVA (significance level α = 0.05) and Dunnett's multi-comparison post-hoc tests (mutations vs. wild-type). * P
    Pgfp V Rs Cloning Vector, supplied by OriGene, used in various techniques. Bioz Stars score: 79/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    83
    OriGene human myotilin
    FOXM1 directly binds to the putative FOXM1 consensus binding sequences located on the <t>CDC25A</t> promoter. (A) FOXM1 binding to CDC25A promoter was confirmed by ChIP-qPCR. Primer set A was designed to span three FOXM1 binding sites, and primer set B, targeting a region of the promoter that did not include the FOXM1 binding sites, was used as the control. Chromatin immunoprecipitations of U2OS cells were prepared using anti-FOXM1 antibody. Semi-quantitative PCR was performed using the primer sets A and B, and the PCR products were detected by electrophoresis (Top). Quantitative PCR was performed using the primer sets A and B, and the binding activity of FOXM1 to CDC25A promoter was evaluated by the fold enrichment method. (B–C) The three FOXM1 binding sites on the CDC25A promoter are functionally redundant (B) Schematic diagram showing the mutations of the three FOXM1 binding sites on the CDC25A promoter. (C) U2OS cells were transfected with pACT-FOXM1 or empty vector control, along with the indicated CDC25A promoter-luciferase reporter construct. Targeted mutation of single FOXM1 binding sites did not significantly affect CDC25A transactivation. Mutations of sites 1 and 2, sites 2 and 3, and sites 1, 2, and 3 significantly reduced the transactivation of the CDC25A-luciferase reporter. Data were normalized to Renilla luciferase activities and are presented as the mean wild-typefold induction over empty vector control ± SD (N = 3). Data were subjected to one-way ANOVA (significance level α = 0.05) and Dunnett's multi-comparison post-hoc tests (mutations vs. wild-type). * P
    Human Myotilin, supplied by OriGene, used in various techniques. Bioz Stars score: 83/100, based on 22 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    78
    Agilent technologies pcmv3tag9
    CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with <t>pCMV3Tag9-CDC25A</t> (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p
    Pcmv3tag9, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 78/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    OriGene cdna template
    CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with <t>pCMV3Tag9-CDC25A</t> (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p
    Cdna Template, supplied by OriGene, used in various techniques. Bioz Stars score: 92/100, based on 38 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    80
    OriGene number rc220744
    CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with <t>pCMV3Tag9-CDC25A</t> (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p
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    OriGene pcmv6 xl5 tdg
    CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with <t>pCMV3Tag9-CDC25A</t> (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p
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    85
    Roche bamhi sites
    CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with <t>pCMV3Tag9-CDC25A</t> (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p
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    OriGene hush pgfp v rs plasmid
    CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with <t>pCMV3Tag9-CDC25A</t> (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p
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    OriGene aff4 cdna
    CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with <t>pCMV3Tag9-CDC25A</t> (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p
    Aff4 Cdna, supplied by OriGene, used in various techniques. Bioz Stars score: 78/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Stratagene bamhi site
    CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with <t>pCMV3Tag9-CDC25A</t> (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p
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    OriGene cd11b cdnas
    CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with <t>pCMV3Tag9-CDC25A</t> (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p
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    Thermo Fisher human keratin 18
    CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with <t>pCMV3Tag9-CDC25A</t> (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p
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    Image Search Results


    Direct visualization of HPV88 on an EtBr-stained gel, after digestion with BamHI, lane 1. Lane M, Gene Ruler 1 kb DNA ladder (Fermentas).

    Journal: International journal of cancer. Journal international du cancer

    Article Title: Cutaneous human papillomavirus 88: Remarkable differences in viral load

    doi: 10.1002/ijc.23115

    Figure Lengend Snippet: Direct visualization of HPV88 on an EtBr-stained gel, after digestion with BamHI, lane 1. Lane M, Gene Ruler 1 kb DNA ladder (Fermentas).

    Article Snippet: Fifty microliters of unamplified extracted DNA from the original tumor of the index patient, containing ~5 × 1010 copies of HPV88, was digested with 20 U of BamHI (Fermentas).

    Techniques: Staining

    FOXM1 directly binds to the putative FOXM1 consensus binding sequences located on the CDC25A promoter. (A) FOXM1 binding to CDC25A promoter was confirmed by ChIP-qPCR. Primer set A was designed to span three FOXM1 binding sites, and primer set B, targeting a region of the promoter that did not include the FOXM1 binding sites, was used as the control. Chromatin immunoprecipitations of U2OS cells were prepared using anti-FOXM1 antibody. Semi-quantitative PCR was performed using the primer sets A and B, and the PCR products were detected by electrophoresis (Top). Quantitative PCR was performed using the primer sets A and B, and the binding activity of FOXM1 to CDC25A promoter was evaluated by the fold enrichment method. (B–C) The three FOXM1 binding sites on the CDC25A promoter are functionally redundant (B) Schematic diagram showing the mutations of the three FOXM1 binding sites on the CDC25A promoter. (C) U2OS cells were transfected with pACT-FOXM1 or empty vector control, along with the indicated CDC25A promoter-luciferase reporter construct. Targeted mutation of single FOXM1 binding sites did not significantly affect CDC25A transactivation. Mutations of sites 1 and 2, sites 2 and 3, and sites 1, 2, and 3 significantly reduced the transactivation of the CDC25A-luciferase reporter. Data were normalized to Renilla luciferase activities and are presented as the mean wild-typefold induction over empty vector control ± SD (N = 3). Data were subjected to one-way ANOVA (significance level α = 0.05) and Dunnett's multi-comparison post-hoc tests (mutations vs. wild-type). * P

    Journal: PLoS ONE

    Article Title: Novel Interactions between FOXM1 and CDC25A Regulate the Cell Cycle

    doi: 10.1371/journal.pone.0051277

    Figure Lengend Snippet: FOXM1 directly binds to the putative FOXM1 consensus binding sequences located on the CDC25A promoter. (A) FOXM1 binding to CDC25A promoter was confirmed by ChIP-qPCR. Primer set A was designed to span three FOXM1 binding sites, and primer set B, targeting a region of the promoter that did not include the FOXM1 binding sites, was used as the control. Chromatin immunoprecipitations of U2OS cells were prepared using anti-FOXM1 antibody. Semi-quantitative PCR was performed using the primer sets A and B, and the PCR products were detected by electrophoresis (Top). Quantitative PCR was performed using the primer sets A and B, and the binding activity of FOXM1 to CDC25A promoter was evaluated by the fold enrichment method. (B–C) The three FOXM1 binding sites on the CDC25A promoter are functionally redundant (B) Schematic diagram showing the mutations of the three FOXM1 binding sites on the CDC25A promoter. (C) U2OS cells were transfected with pACT-FOXM1 or empty vector control, along with the indicated CDC25A promoter-luciferase reporter construct. Targeted mutation of single FOXM1 binding sites did not significantly affect CDC25A transactivation. Mutations of sites 1 and 2, sites 2 and 3, and sites 1, 2, and 3 significantly reduced the transactivation of the CDC25A-luciferase reporter. Data were normalized to Renilla luciferase activities and are presented as the mean wild-typefold induction over empty vector control ± SD (N = 3). Data were subjected to one-way ANOVA (significance level α = 0.05) and Dunnett's multi-comparison post-hoc tests (mutations vs. wild-type). * P

    Article Snippet: The CDC25A plasmid was purchased from Origene and subcloned in-frame in the pACT expression vector at the BamHI and XbaI restriction sites or the pCMV3Tag9 (Agilent, La Jolla, CA) expression vector at the BamHI and XhoI restriction sites.

    Techniques: Binding Assay, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Polymerase Chain Reaction, Electrophoresis, Activity Assay, Transfection, Plasmid Preparation, Luciferase, Construct, Mutagenesis

    CDC25A phosphatase enzyme activity is required for the CDC25A-activated FOXM1 transcriptional activity and protein-protein interaction between CDC25A and FOXM1. (A) CDC25A phosphatase enzyme activity is required for the CDC25A-activated FOXM1 transcriptional activity. The C431 site in CDC25A protein was changed via site-directed mutagenesis to a serine to create a phosphatase-dead mutant. U2OS cells were co-transfected with pACT-FOXM1 and/or pACT-CDC25A (pACT-CDC25A C431S), along with the pGL3-6×FOXM1-Luc plasmids containing 6 FOXM1 DNA binding sequences. pRL-SV40, a Renilla luciferase reporter vector, was used as the control. Data were normalized to Renilla luciferase activities and are presented as the mean ± SD (N = 5). *** p

    Journal: PLoS ONE

    Article Title: Novel Interactions between FOXM1 and CDC25A Regulate the Cell Cycle

    doi: 10.1371/journal.pone.0051277

    Figure Lengend Snippet: CDC25A phosphatase enzyme activity is required for the CDC25A-activated FOXM1 transcriptional activity and protein-protein interaction between CDC25A and FOXM1. (A) CDC25A phosphatase enzyme activity is required for the CDC25A-activated FOXM1 transcriptional activity. The C431 site in CDC25A protein was changed via site-directed mutagenesis to a serine to create a phosphatase-dead mutant. U2OS cells were co-transfected with pACT-FOXM1 and/or pACT-CDC25A (pACT-CDC25A C431S), along with the pGL3-6×FOXM1-Luc plasmids containing 6 FOXM1 DNA binding sequences. pRL-SV40, a Renilla luciferase reporter vector, was used as the control. Data were normalized to Renilla luciferase activities and are presented as the mean ± SD (N = 5). *** p

    Article Snippet: The CDC25A plasmid was purchased from Origene and subcloned in-frame in the pACT expression vector at the BamHI and XbaI restriction sites or the pCMV3Tag9 (Agilent, La Jolla, CA) expression vector at the BamHI and XhoI restriction sites.

    Techniques: Activity Assay, Mutagenesis, Transfection, Binding Assay, Luciferase, Plasmid Preparation

    CDC25A regulated FOXM1 transcriptional activity through CDK1-mediated phosphorylation sites. (A) CDC25A activated the transcriptional activity of FOXM1 in U2OS cells. U2OS cells were co-transfected with pACT-FOXM1 or/and pACT-CDC25A, along with the pGL3-6×FOXM1-Luc plasmids containing 6 FOXM1 DNA binding sequences, and pRL-SV40, a Renilla luciferase reporter vector was used as the control. Data were normalized to Renilla luciferase activities and are presented as the mean ± SD (N = 3). *** p

    Journal: PLoS ONE

    Article Title: Novel Interactions between FOXM1 and CDC25A Regulate the Cell Cycle

    doi: 10.1371/journal.pone.0051277

    Figure Lengend Snippet: CDC25A regulated FOXM1 transcriptional activity through CDK1-mediated phosphorylation sites. (A) CDC25A activated the transcriptional activity of FOXM1 in U2OS cells. U2OS cells were co-transfected with pACT-FOXM1 or/and pACT-CDC25A, along with the pGL3-6×FOXM1-Luc plasmids containing 6 FOXM1 DNA binding sequences, and pRL-SV40, a Renilla luciferase reporter vector was used as the control. Data were normalized to Renilla luciferase activities and are presented as the mean ± SD (N = 3). *** p

    Article Snippet: The CDC25A plasmid was purchased from Origene and subcloned in-frame in the pACT expression vector at the BamHI and XbaI restriction sites or the pCMV3Tag9 (Agilent, La Jolla, CA) expression vector at the BamHI and XhoI restriction sites.

    Techniques: Activity Assay, Transfection, Binding Assay, Luciferase, Plasmid Preparation

    FOXM1 regulates the gene transcription of CDC25A by direct DNA binding as well as the E2F pathway. (A) Schematic diagram of CDC25A promoter showing the 3′ distal end containing two E2F binding sites, the 5′ proximal end including three putative FOXM1 binding sites and excluding E2F binding sites, and a 224 bp fragment containing the three putative FOXM1 binding sites. (B) FOXM1 increased the transactivation of full-length and truncated CDC25A promoter constructs, indicating both direct and indirect FOXM1 induction. U2OS cells were co-transfected with pACT-FOXM1 or pACT empty plasmid and the reporter plasmids containing full length and truncated CDC25A promoters. pRL-SV40, a Renilla luciferase report vector, was co-transfected as the control. Data were normalized to Renilla luciferase activities and are presented as the mean ± SD (N = 5). ***, p

    Journal: PLoS ONE

    Article Title: Novel Interactions between FOXM1 and CDC25A Regulate the Cell Cycle

    doi: 10.1371/journal.pone.0051277

    Figure Lengend Snippet: FOXM1 regulates the gene transcription of CDC25A by direct DNA binding as well as the E2F pathway. (A) Schematic diagram of CDC25A promoter showing the 3′ distal end containing two E2F binding sites, the 5′ proximal end including three putative FOXM1 binding sites and excluding E2F binding sites, and a 224 bp fragment containing the three putative FOXM1 binding sites. (B) FOXM1 increased the transactivation of full-length and truncated CDC25A promoter constructs, indicating both direct and indirect FOXM1 induction. U2OS cells were co-transfected with pACT-FOXM1 or pACT empty plasmid and the reporter plasmids containing full length and truncated CDC25A promoters. pRL-SV40, a Renilla luciferase report vector, was co-transfected as the control. Data were normalized to Renilla luciferase activities and are presented as the mean ± SD (N = 5). ***, p

    Article Snippet: The CDC25A plasmid was purchased from Origene and subcloned in-frame in the pACT expression vector at the BamHI and XbaI restriction sites or the pCMV3Tag9 (Agilent, La Jolla, CA) expression vector at the BamHI and XhoI restriction sites.

    Techniques: Binding Assay, Construct, Transfection, Plasmid Preparation, Luciferase

    Coordinated expression of FOXM1 and CDC25A during the cell cycle progression. (A) Representative flow cytometry analysis shows cell cycle progression in U2OS cells. U2OS were synchronized with nocodazole for 16 h and then stimulated to re-enter the cell cycle by addition of medium containing 10% fetal bovine serum. Asynchronous cells (Asynch) were included as controls. Cell cycle was analyzed by flow cytometry at release (16 h Noc) and at 1, 3, 6, 12, and 18 h after release. (B) Mean percentage (± SD) of cells in each phase of the cell cycle following 16 h nocodazole treatment and after 1, 3, 6, 12, 18 h release (N = 3). Asynchronous (Asynch) cells were included as controls. The analysis indicates that the cells at early, middle and late G1 phase are at 1, 3 and 6 h respectively, S phase is maximal at 12 h while G2/M phase occurs at 16 h Noc and 18 h after release. (C) Cells at the continuous cell cycle phases were collected and processed for western blotting analysis with antibodies against FOXM1, CDC25A, CDK1 and CDK2. FOXM1 expression increased as cells progressed from G1 through S and into G2/M, and degraded when cells exited to G2/M (1 h). CDC25A and CDK1 exhibited a similar expression profile. CDK2 expression levels were relatively constant throughout the cell cycle. β-Actin expression was used as a loading control.

    Journal: PLoS ONE

    Article Title: Novel Interactions between FOXM1 and CDC25A Regulate the Cell Cycle

    doi: 10.1371/journal.pone.0051277

    Figure Lengend Snippet: Coordinated expression of FOXM1 and CDC25A during the cell cycle progression. (A) Representative flow cytometry analysis shows cell cycle progression in U2OS cells. U2OS were synchronized with nocodazole for 16 h and then stimulated to re-enter the cell cycle by addition of medium containing 10% fetal bovine serum. Asynchronous cells (Asynch) were included as controls. Cell cycle was analyzed by flow cytometry at release (16 h Noc) and at 1, 3, 6, 12, and 18 h after release. (B) Mean percentage (± SD) of cells in each phase of the cell cycle following 16 h nocodazole treatment and after 1, 3, 6, 12, 18 h release (N = 3). Asynchronous (Asynch) cells were included as controls. The analysis indicates that the cells at early, middle and late G1 phase are at 1, 3 and 6 h respectively, S phase is maximal at 12 h while G2/M phase occurs at 16 h Noc and 18 h after release. (C) Cells at the continuous cell cycle phases were collected and processed for western blotting analysis with antibodies against FOXM1, CDC25A, CDK1 and CDK2. FOXM1 expression increased as cells progressed from G1 through S and into G2/M, and degraded when cells exited to G2/M (1 h). CDC25A and CDK1 exhibited a similar expression profile. CDK2 expression levels were relatively constant throughout the cell cycle. β-Actin expression was used as a loading control.

    Article Snippet: The CDC25A plasmid was purchased from Origene and subcloned in-frame in the pACT expression vector at the BamHI and XbaI restriction sites or the pCMV3Tag9 (Agilent, La Jolla, CA) expression vector at the BamHI and XhoI restriction sites.

    Techniques: Expressing, Flow Cytometry, Cytometry, Western Blot

    CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with pCMV3Tag9-CDC25A (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p

    Journal: PLoS ONE

    Article Title: Novel Interactions between FOXM1 and CDC25A Regulate the Cell Cycle

    doi: 10.1371/journal.pone.0051277

    Figure Lengend Snippet: CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with pCMV3Tag9-CDC25A (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p

    Article Snippet: The CDC25A plasmid was purchased from Origene and subcloned in-frame in the pACT expression vector at the BamHI and XbaI restriction sites or the pCMV3Tag9 (Agilent, La Jolla, CA) expression vector at the BamHI and XhoI restriction sites.

    Techniques: Activity Assay, Immunoprecipitation, Transfection, Plasmid Preparation, Polyacrylamide Gel Electrophoresis, Western Blot, Incubation, Construct, Two Hybrid Assay, Luciferase, Expressing

    The FOXM1 transcription factor regulates CDC25A gene expression. (A) FOXM1 increased the transactivation of CDC25A promoter. U2OS cells were co-transfected with pGL3-CDC25A promoter reporter along with increasing doses of pACT-FOXM1 for 48 h. pACT vector was used as the empty vector, and pRL-SV40 expressing Renilla luciferase was used as an internal control. Data represent the mean ± SD (N = 3). *, p

    Journal: PLoS ONE

    Article Title: Novel Interactions between FOXM1 and CDC25A Regulate the Cell Cycle

    doi: 10.1371/journal.pone.0051277

    Figure Lengend Snippet: The FOXM1 transcription factor regulates CDC25A gene expression. (A) FOXM1 increased the transactivation of CDC25A promoter. U2OS cells were co-transfected with pGL3-CDC25A promoter reporter along with increasing doses of pACT-FOXM1 for 48 h. pACT vector was used as the empty vector, and pRL-SV40 expressing Renilla luciferase was used as an internal control. Data represent the mean ± SD (N = 3). *, p

    Article Snippet: The CDC25A plasmid was purchased from Origene and subcloned in-frame in the pACT expression vector at the BamHI and XbaI restriction sites or the pCMV3Tag9 (Agilent, La Jolla, CA) expression vector at the BamHI and XhoI restriction sites.

    Techniques: Expressing, Transfection, Plasmid Preparation, Luciferase

    CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with pCMV3Tag9-CDC25A (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p

    Journal: PLoS ONE

    Article Title: Novel Interactions between FOXM1 and CDC25A Regulate the Cell Cycle

    doi: 10.1371/journal.pone.0051277

    Figure Lengend Snippet: CDC25A phosphatase activates FOXM1 transcriptional activity by direct protein-protein interaction. (A) CDC25A and FOXM1 proteins physically interact. The protein-protein interaction of FOXM1 and CDC25A was confirmed by co-immunoprecipitation. HEK293T cells were co-transfected with p3×FLAG-CMV-14-FOXM1 (FOXM1-3×FLAG) or the empty vector control p3×FLAG-CMV-14, along with pCMV3Tag9-CDC25A (CDC25A-3×MYC) or the empty vector control (pCMV3Tag9). Lysates were immunoprecipitated with anti-FLAG agarose resin, separated by PAGE, and electroblotted to PVDF. Western blot analysis showed that CDC25A-3×MYC co-immunoprecipitates with FOXM1-3×FLAG, supporting the hypothesis that these two expressed proteins interact. (B) The native protein interaction between FOXM1 and CDC25A was detected by co-immunoprecipitation. 6×10 6 U2OS cells were lysed in MPER buffer containing 1× protease/phosphatase inhibitors. Whole cell lysates were pre-cleared with protein G sepharose and normal rabbit IgG overnight at 4°C with end-over-end mixing. The cell lysates were incubated with anti-CDC25A antibody overnight at 4°C and then with protein G sepharose for 1 h at 4°C. The resin was washed three times, and the eluted protein complex was separated by PAGE and analyzed using the anti-FOXM1 antibody by Western blotting. (C) Schematic diagram showing FOXM1 deletion constructs used to identify FOXM1 domains critical to the interaction with CDC25A. Sequences encoding the specific FOXM1 deletion constructs were subcloned in pBIND. (D) A mammalian two-hybrid assay reveals a critical role for the FOXM1 C-terminus in interactions with CDC25A. pBIND construct (FOXM1, deletion, or empty) was co-transfected with pACT construct (CDC25A or empty) and pG5luc reporter. Data represent the mean ± SD, normalized to Renilla luciferase activities (N = 3). Co-expression of the C-terminal FOXM1 and CDC25A revealed a robust interaction. In a construct lacking the C-terminus, the interaction between FOXM1 and CDC25A was significantly diminished. *** p

    Article Snippet: The CDC25A plasmid was purchased from Origene and subcloned in-frame in the pACT expression vector at the BamHI and XbaI restriction sites or the pCMV3Tag9 (Agilent, La Jolla, CA) expression vector at the BamHI and XhoI restriction sites.

    Techniques: Activity Assay, Immunoprecipitation, Transfection, Plasmid Preparation, Polyacrylamide Gel Electrophoresis, Western Blot, Incubation, Construct, Two Hybrid Assay, Luciferase, Expressing