human bmp4 cdna expression Search Results


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  • 99
    Thermo Fisher gene exp bmp4 mm00432087 m1
    Transient Tbx6 Expression Induced Mesoderm and Cardiovascular Lineages in Mouse ESCs (A) Schematic representation of the lentiviral construct for Dox-inducible Tbx6 expression. (B) Clonal expansion of the iTbx6 T-GFP mouse ESCs was confirmed by hKO expression. (C) The mRNA expression of Tbx6 in iTbx6 T-GFP mESCs with or without Dox administration was determined by qRT-PCR (n = 3, independent experiments). Data were normalized to the values in No Dox. (D) Scheme depicting the protocol used to evaluate the effects of Tbx6 expression on mesoderm induction. ABV indicates Activin A, <t>BMP4,</t> and VEGF. . (H) The protocol used to evaluate the effects of the duration of Tbx6 expression on cardiac differentiation. . (K) iTbx6 T-GFP mESCs were differentiated into CMs, SMCs, and ECs with Dox administration for 3 days. Representative images are shown. (L) The mRNA expression for CM, SMC, and EC genes in iTbx6 T-GFP mESCs with (red, Dox on D0–3) or without Dox (white, No Dox) in the absence of cytokines (n = 3, independent experiments). ABV (black) is cytokine-based cardiac differentiation. Data were normalized to the values in No Dox. .
    Gene Exp Bmp4 Mm00432087 M1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 135 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore tri reagent
    Transient Tbx6 Expression Induced Mesoderm and Cardiovascular Lineages in Mouse ESCs (A) Schematic representation of the lentiviral construct for Dox-inducible Tbx6 expression. (B) Clonal expansion of the iTbx6 T-GFP mouse ESCs was confirmed by hKO expression. (C) The mRNA expression of Tbx6 in iTbx6 T-GFP mESCs with or without Dox administration was determined by qRT-PCR (n = 3, independent experiments). Data were normalized to the values in No Dox. (D) Scheme depicting the protocol used to evaluate the effects of Tbx6 expression on mesoderm induction. ABV indicates Activin A, <t>BMP4,</t> and VEGF. . (H) The protocol used to evaluate the effects of the duration of Tbx6 expression on cardiac differentiation. . (K) iTbx6 T-GFP mESCs were differentiated into CMs, SMCs, and ECs with Dox administration for 3 days. Representative images are shown. (L) The mRNA expression for CM, SMC, and EC genes in iTbx6 T-GFP mESCs with (red, Dox on D0–3) or without Dox (white, No Dox) in the absence of cytokines (n = 3, independent experiments). ABV (black) is cytokine-based cardiac differentiation. Data were normalized to the values in No Dox. .
    Tri Reagent, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 45305 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    Wyeth Biopharma human bmp4
    Retinoic acid (RA) and <t>BMP4</t> counteract each other's inhibition of skeletal myogenesis or cardiomyogenesis . P19 cells were mixed with P19[BMP4] or P19[control] cells in the presence of 1% dimethylsulfoxide (DMSO), with or without RA. Panel I : P19[BMP4] cultures treated with RA were fixed on day 9 for immunofluorescence with MF20 antibody (A, B) and counter stained with Hoechst dye (C, D). Magnification is 160x. Panel II : Skeletal myogenesis was quantified for each condition by counting the number of myosin heavy chain +ve bipolar skeletal myocytes, expressed as the percentage of total cells (white bars) and their standard errors ( n = 3). MyoD transcript levels (black bars) were quantified by quantitative polymerase chain reaction and expressed relative to control cultures ( n = 2), * P
    Human Bmp4, supplied by Wyeth Biopharma, used in various techniques. Bioz Stars score: 89/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher human kidney cdna library
    Retinoic acid (RA) and <t>BMP4</t> counteract each other's inhibition of skeletal myogenesis or cardiomyogenesis . P19 cells were mixed with P19[BMP4] or P19[control] cells in the presence of 1% dimethylsulfoxide (DMSO), with or without RA. Panel I : P19[BMP4] cultures treated with RA were fixed on day 9 for immunofluorescence with MF20 antibody (A, B) and counter stained with Hoechst dye (C, D). Magnification is 160x. Panel II : Skeletal myogenesis was quantified for each condition by counting the number of myosin heavy chain +ve bipolar skeletal myocytes, expressed as the percentage of total cells (white bars) and their standard errors ( n = 3). MyoD transcript levels (black bars) were quantified by quantitative polymerase chain reaction and expressed relative to control cultures ( n = 2), * P
    Human Kidney Cdna Library, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 91/100, based on 92 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems human bmp4
    TWSG1 inhibit up-regulation of hepcidin expression by BMP2/4 in human hepatoma cell line . (A) Relative hepcidin expression as a dosed response to BMP2 (□) and <t>BMP4</t> (■) are shown. Final BMP concentrations are on the x-axis, and relative
    Human Bmp4, supplied by R&D Systems, used in various techniques. Bioz Stars score: 96/100, based on 174 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems bmp4
    Lung epithelial branching morphogenesis was severely impaired in Apc CKO embryos. a Lung epithelial branches at E13.5 were visualized by whole mount E-cadherin immunofluorescence staining. b Altered expression of Fgf10 and <t>Bmp4</t> in Apc CKO lung at E11.5 was detected by real-time PCR. * P
    Bmp4, supplied by R&D Systems, used in various techniques. Bioz Stars score: 98/100, based on 2875 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Qiagen fastlane cell cdna kit
    Lung epithelial branching morphogenesis was severely impaired in Apc CKO embryos. a Lung epithelial branches at E13.5 were visualized by whole mount E-cadherin immunofluorescence staining. b Altered expression of Fgf10 and <t>Bmp4</t> in Apc CKO lung at E11.5 was detected by real-time PCR. * P
    Fastlane Cell Cdna Kit, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 369 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher gene exp bmp4 hs00370078 m1
    Lung epithelial branching morphogenesis was severely impaired in Apc CKO embryos. a Lung epithelial branches at E13.5 were visualized by whole mount E-cadherin immunofluorescence staining. b Altered expression of Fgf10 and <t>Bmp4</t> in Apc CKO lung at E11.5 was detected by real-time PCR. * P
    Gene Exp Bmp4 Hs00370078 M1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 91/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    OriGene wildtype cdna
    Lung epithelial branching morphogenesis was severely impaired in Apc CKO embryos. a Lung epithelial branches at E13.5 were visualized by whole mount E-cadherin immunofluorescence staining. b Altered expression of Fgf10 and <t>Bmp4</t> in Apc CKO lung at E11.5 was detected by real-time PCR. * P
    Wildtype Cdna, supplied by OriGene, used in various techniques. Bioz Stars score: 93/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    InvivoGen vegf165 cdna
    Lung epithelial branching morphogenesis was severely impaired in Apc CKO embryos. a Lung epithelial branches at E13.5 were visualized by whole mount E-cadherin immunofluorescence staining. b Altered expression of Fgf10 and <t>Bmp4</t> in Apc CKO lung at E11.5 was detected by real-time PCR. * P
    Vegf165 Cdna, supplied by InvivoGen, used in various techniques. Bioz Stars score: 85/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    OriGene ddk tagged expression vector
    Lung epithelial branching morphogenesis was severely impaired in Apc CKO embryos. a Lung epithelial branches at E13.5 were visualized by whole mount E-cadherin immunofluorescence staining. b Altered expression of Fgf10 and <t>Bmp4</t> in Apc CKO lung at E11.5 was detected by real-time PCR. * P
    Ddk Tagged Expression Vector, supplied by OriGene, used in various techniques. Bioz Stars score: 93/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher pcdna3 1zeo man expression vector
    Lung epithelial branching morphogenesis was severely impaired in Apc CKO embryos. a Lung epithelial branches at E13.5 were visualized by whole mount E-cadherin immunofluorescence staining. b Altered expression of Fgf10 and <t>Bmp4</t> in Apc CKO lung at E11.5 was detected by real-time PCR. * P
    Pcdna3 1zeo Man Expression Vector, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 98/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    PeproTech human twsg1
    <t>Twsg1</t> dose response of hepcidin levels in primary murine hepatocytes . In primary hepatocytes from 4- to 6-week-old C57Bl/6 mice, dosed increased Twsg1 (x-axis) was added, and hepcidin (y-axis; hamp1) mRNA was quantified. * P
    Human Twsg1, supplied by PeproTech, used in various techniques. Bioz Stars score: 85/100, based on 10 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Qiagen human common cytokines
    Expression of primary dysmenorrhea-related genes by quantitative RT-PCR array on the seventh day before menstruation, and the first and fifth days of menstruation. Compared with the unaffected control group, the primary dysmenorrhea group has the relatively low expression of genes ( BMP6 , GDF5 , GDF11 , NODAL , IL1F5 , IL11 and MSTN ), and high expression of pro-inflammatory <t>cytokines</t> ( IL1B, TNF, IL6, and IL8 ).
    Human Common Cytokines, supplied by Qiagen, used in various techniques. Bioz Stars score: 85/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam rabbit anti human gata4
    Expression of cardiac markers in somite cells from embryos lacking Myo/Nog cells Somite cultures were prepared from untreated and G8 and complement treated stage 12–13 embryos. Somite cells from untreated embryos formed sarcomeric myosin+, elongated skeletal myotubes (A). Cultures from ablated embryos contained <t>GATA4+</t> and cardiac troponin I+ (CTpI) cells (B). Some cardiomyocytes contained striations (arrows in B). CTpI+ cells in somite cultures resembled cultured cardiomyocytes from the stage 12 heart (C). In vivo , the somites of G8 ablated (G, H) but not untreated (D, E) stage 12–16 (D, G) and stage 25–26 embryos (E and H) contained GATA4+ and CTpI+ cells. Cardiac troponin T (CTpT) was expressed in stage 25–26 ablated (I) but not untreated embryos (F). Bar, 9 µm.
    Rabbit Anti Human Gata4, supplied by Abcam, used in various techniques. Bioz Stars score: 85/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    R&D Systems bmp2
    BMP and Noggin influence neural crest-derived migration and neurite patterning in E11.5 mouse midgut Gut explants were grown in culture with the cut end of the midgut placed onto filter paper to allow outgrowth (see diagram G). Migration and neurite extension from the midgut was examined after 48 hours in culture with GDNF (A, A′, A″), GDNF plus noggin (B, B′, B″), GDNF plus anti-BMP4 blocking antibody (C, C′, C″), GDNF plus <t>BMP2</t> (D, D′, D″), or GDNF plus BMP4 (E, E′, E″). Immunohistochemistry was performed for TuJ1 (A–E) or Ret (A′–E′). Merged images (A″ – E″). (H) The schematic shows how the extent of migration from the edge of the explant was measured. Seven radiating lines were drawn in standard positions extending from the edge of the explant. The position of the most distant Ret+ cell along these lines was recorded. (F) Mean distance that Ret+ cells migrated from the edge of the explant confirms increased migration with GDNF plus noggin or GDNF plus anti-BMP4 blocking antibody, and reduced migration with GDNF plus BMP4 (n = 12 explants) from 6 separate experiments. Scale bar = 100 μm. (I–L) To confirm the location of Ret+ cells that migrated from the explants, cell nuclei were visualized using Hoechst 33342 dye. Low magnification images of an explant grown in GDNF plus noggin are shown after immunohistochemistry for TuJ1 (I), Ret (J) and Hoechst 33342 (K) with an additional merged image (L). The white box in (L) marks the region shown in (N). (M–P) Show higher magnification images of cells and neurites at the edge of the outgrowth from explants after growth in GDNF (M), GDNF plus noggin (N), GDNF plus BMP4 blocking antibody (O) or GDNF plus BMP4 (P). These images demonstrate cell bodies at the tips of the furthest neurites and highlight BMP4 induced neurite fasciculation (P). * P
    Bmp2, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 974 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology rabbit anti human smad4
    Effects of Myo/Nog cell ablation on the expression of regulators of BMP signaling in the stage 2 embryo Untreated embryos and G8 or D4/complement treated stage X–XII embryos were incubated to stage 2 and double labeled with antibodies. The colors of the fluorescent tags are indicated in each panel. The photograph of the stage 2 embryo (A) illustrates the regions shown in the fluorescence micrographs. Overlap of red and green appears yellow/white in merged images. After ablating G8+ cells in the stage X–XII epiblast, neither G8 (B, C, E–I, K–N), noggin (NogP) (B) nor follistatin (Fol) (C) was detected in the stage 2 embryo. Ablation with the D4 MAb did not affect follistatin expression (D). Staining for chordin (Chor) was reduced along Koller’s sickle (ks) in G8 ablated embryos (E). In untreated stage 2 embryos, BMP4+ cells surrounded Koller’s sickle (F) and BMP2 was present below Koller’s sickle (bks) (G). P-Smad1/5/8 was detected only below Koller’s sickle (H) and did not overlap with G8+ cells in the posterior/medial epiblast (pme) (I). Cytoplasmic staining for <t>Smad4</t> was present in the pme (J). In G8 ablated embryos, staining for BMP4 was increased in the pme (K), BMP2 remained below Koller’s sickle (L) and p-Smad1/5/8 was expanded into the pme (M) and central epiblast (ce) (N). P-Smad1/5/8 was not present above Koller’s sickle in embryos ablated with the D4 MAb (O). Bar, 1,250 µm in A; 9 µm in B–T.
    Rabbit Anti Human Smad4, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 85/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology goat anti human follistatin
    Effects of reintroducing Myo/Nog cells into ablated embryos on regulators of BMP signaling in the stage 2 epiblast G8+ cells isolated from stage X–XII embryos were injected into the epiblast after the indigenous population of G8+ cells was ablated with complement. Embryos were grown to stage 2 and double labeled with antibodies. The photograph of the stage 2 embryo (A) illustrates the regions shown in the fluorescence micrographs. The colors of the fluorescent tags are indicated in each panel. Overlap of red and green appears yellow in merged images. Implanted G8+ cells synthesized noggin in the posterior/medial epiblast (pme) (B) and in the anterior portion of the developing primitive streak (as) (C), restored expression of <t>follistatin</t> in the posterior/lateral epiblast (ple) (D) and inhibited p-Smad1/5/8 accumulation in the epiblast in the pme (E). P-Smad1/5/8 was present below Koller’s sickle (bks) (F). Bar, 900 µm in A; 9 µm in B–F.
    Goat Anti Human Follistatin, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 85/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher sequence specific taqman gene expression assays
    Effects of reintroducing Myo/Nog cells into ablated embryos on regulators of BMP signaling in the stage 2 epiblast G8+ cells isolated from stage X–XII embryos were injected into the epiblast after the indigenous population of G8+ cells was ablated with complement. Embryos were grown to stage 2 and double labeled with antibodies. The photograph of the stage 2 embryo (A) illustrates the regions shown in the fluorescence micrographs. The colors of the fluorescent tags are indicated in each panel. Overlap of red and green appears yellow in merged images. Implanted G8+ cells synthesized noggin in the posterior/medial epiblast (pme) (B) and in the anterior portion of the developing primitive streak (as) (C), restored expression of <t>follistatin</t> in the posterior/lateral epiblast (ple) (D) and inhibited p-Smad1/5/8 accumulation in the epiblast in the pme (E). P-Smad1/5/8 was present below Koller’s sickle (bks) (F). Bar, 900 µm in A; 9 µm in B–F.
    Sequence Specific Taqman Gene Expression Assays, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 26 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems recombinant chordin like 1 protein
    Effect of <t>Chordin-like</t> 1 and SB203580 on expression of p21 Cip1/WAF1 , p53, phospho-Rb, and Apo J. A , Western blot analysis of the protein levels of p21 Cip1/WAF1 , p53, and phospho-Rb ( pRb ) in oxidant-treated ARPE-19 cells, with or without Chordin-like
    Recombinant Chordin Like 1 Protein, supplied by R&D Systems, 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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    Agilent technologies whole human genome 44k microarrays
    Effect of <t>Chordin-like</t> 1 and SB203580 on expression of p21 Cip1/WAF1 , p53, phospho-Rb, and Apo J. A , Western blot analysis of the protein levels of p21 Cip1/WAF1 , p53, and phospho-Rb ( pRb ) in oxidant-treated ARPE-19 cells, with or without Chordin-like
    Whole Human Genome 44k Microarrays, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 85/100, based on 30 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher human genome u133 plus 2 0
    Examples of cell type independent, human ES cell and B cell specific Myc targets. ( A ) Gene expression levels (at log2 scale) of FBL , LIN28 and BLMH in the absence and presence of Myc induction in P493-6 B cells (measured by Affymetrix Exon arrays and Affymetrix <t>U133</t> Plus 2.0 arrays), and log2 gene expression fold changes between hESC and trophoblasts (measured by Agilent microarrays) are shown. Error bars correspond to standard deviations of replicate samples. ( B ) ChIP-chip binding signals in human P493-6 B cells and H9 ES cells, and ENCODE ChIP-seq binding signals in three human cancer cell lines. For ChIP-chip, TileMap moving average statistic [62] m was computed for each probe using normalized log2 probe intensities, and 2 m was displayed as the intensity measure. For ChIP-seq, a 100 bp sliding window was used to scan the genome. Read count in each window was shown at a 25 bp step size. E-box motifs CACGTG (black) and CANNTG (red) were mapped to peak regions and are also shown. ( C ) ChIP-chip [34] and ChIP-seq [40] binding signals in mouse ES cells.
    Human Genome U133 Plus 2 0, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 346 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher ptripz shp21
    Examples of cell type independent, human ES cell and B cell specific Myc targets. ( A ) Gene expression levels (at log2 scale) of FBL , LIN28 and BLMH in the absence and presence of Myc induction in P493-6 B cells (measured by Affymetrix Exon arrays and Affymetrix <t>U133</t> Plus 2.0 arrays), and log2 gene expression fold changes between hESC and trophoblasts (measured by Agilent microarrays) are shown. Error bars correspond to standard deviations of replicate samples. ( B ) ChIP-chip binding signals in human P493-6 B cells and H9 ES cells, and ENCODE ChIP-seq binding signals in three human cancer cell lines. For ChIP-chip, TileMap moving average statistic [62] m was computed for each probe using normalized log2 probe intensities, and 2 m was displayed as the intensity measure. For ChIP-seq, a 100 bp sliding window was used to scan the genome. Read count in each window was shown at a 25 bp step size. E-box motifs CACGTG (black) and CANNTG (red) were mapped to peak regions and are also shown. ( C ) ChIP-chip [34] and ChIP-seq [40] binding signals in mouse ES cells.
    Ptripz Shp21, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 88/100, based on 28 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher phusion site directed mutagenesis kit
    Examples of cell type independent, human ES cell and B cell specific Myc targets. ( A ) Gene expression levels (at log2 scale) of FBL , LIN28 and BLMH in the absence and presence of Myc induction in P493-6 B cells (measured by Affymetrix Exon arrays and Affymetrix <t>U133</t> Plus 2.0 arrays), and log2 gene expression fold changes between hESC and trophoblasts (measured by Agilent microarrays) are shown. Error bars correspond to standard deviations of replicate samples. ( B ) ChIP-chip binding signals in human P493-6 B cells and H9 ES cells, and ENCODE ChIP-seq binding signals in three human cancer cell lines. For ChIP-chip, TileMap moving average statistic [62] m was computed for each probe using normalized log2 probe intensities, and 2 m was displayed as the intensity measure. For ChIP-seq, a 100 bp sliding window was used to scan the genome. Read count in each window was shown at a 25 bp step size. E-box motifs CACGTG (black) and CANNTG (red) were mapped to peak regions and are also shown. ( C ) ChIP-chip [34] and ChIP-seq [40] binding signals in mouse ES cells.
    Phusion Site Directed Mutagenesis Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 3329 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Transient Tbx6 Expression Induced Mesoderm and Cardiovascular Lineages in Mouse ESCs (A) Schematic representation of the lentiviral construct for Dox-inducible Tbx6 expression. (B) Clonal expansion of the iTbx6 T-GFP mouse ESCs was confirmed by hKO expression. (C) The mRNA expression of Tbx6 in iTbx6 T-GFP mESCs with or without Dox administration was determined by qRT-PCR (n = 3, independent experiments). Data were normalized to the values in No Dox. (D) Scheme depicting the protocol used to evaluate the effects of Tbx6 expression on mesoderm induction. ABV indicates Activin A, BMP4, and VEGF. . (H) The protocol used to evaluate the effects of the duration of Tbx6 expression on cardiac differentiation. . (K) iTbx6 T-GFP mESCs were differentiated into CMs, SMCs, and ECs with Dox administration for 3 days. Representative images are shown. (L) The mRNA expression for CM, SMC, and EC genes in iTbx6 T-GFP mESCs with (red, Dox on D0–3) or without Dox (white, No Dox) in the absence of cytokines (n = 3, independent experiments). ABV (black) is cytokine-based cardiac differentiation. Data were normalized to the values in No Dox. .

    Journal: Cell stem cell

    Article Title: Tbx6 Induces Nascent Mesoderm from Pluripotent Stem Cells and Temporally Controls Cardiac versus Somite Lineage Diversification

    doi: 10.1016/j.stem.2018.07.001

    Figure Lengend Snippet: Transient Tbx6 Expression Induced Mesoderm and Cardiovascular Lineages in Mouse ESCs (A) Schematic representation of the lentiviral construct for Dox-inducible Tbx6 expression. (B) Clonal expansion of the iTbx6 T-GFP mouse ESCs was confirmed by hKO expression. (C) The mRNA expression of Tbx6 in iTbx6 T-GFP mESCs with or without Dox administration was determined by qRT-PCR (n = 3, independent experiments). Data were normalized to the values in No Dox. (D) Scheme depicting the protocol used to evaluate the effects of Tbx6 expression on mesoderm induction. ABV indicates Activin A, BMP4, and VEGF. . (H) The protocol used to evaluate the effects of the duration of Tbx6 expression on cardiac differentiation. . (K) iTbx6 T-GFP mESCs were differentiated into CMs, SMCs, and ECs with Dox administration for 3 days. Representative images are shown. (L) The mRNA expression for CM, SMC, and EC genes in iTbx6 T-GFP mESCs with (red, Dox on D0–3) or without Dox (white, No Dox) in the absence of cytokines (n = 3, independent experiments). ABV (black) is cytokine-based cardiac differentiation. Data were normalized to the values in No Dox. .

    Article Snippet: Mouse gene expression assays used the following probes: Acta1 (Mm808218_g1), Actn2 (Mm00473657_m1), Bmp4 (Mm00432087_m1), Cdx2 (Mm01212280_m1), Col2a1 (Mm01309565_m1), Eomes (Mm01351985_m1), Flk1 (Mm01222431 m1), Gapdh (Mm99999915_g1), Gsc (Mm00650681_g1), Isl1 (Mm00517585_m1), Lhx1 (Mm01297482_m1), Meox1 (Mm00440285_m1), Mesp1 (Mm00801883_g1), Msgn1 (Mm00490407_s1), Msx1 (Mm00440330_m1), Myf5 (Mm00435125_m1), Myh3 (Mm01332463_m1), Myh6 (Mm00440354_m1), Myh11 (Mm00443013_m1), Myog (Mm00446194_m1), Nkx2.5 (Mm00657783_m1), Nodal ( Mm00443040_m1), Pax3 (Mm00435491_m1), Pax6 (Mm00443081_m1), Pax7 (Mm01354484_m1), Pdgfr a (Mm00440701_m1), Pecam1 (Mm 01242584_m1), Sox1 (Mm00486299_s1), Sox2 (Mm03053810_s1), Sox9 (Mm00448840_m1), Sox17 (Mm00488363_m1), T (Mm00436877_m1), Tbx6 (Mm01278677_m1), Tcf15 (Mm00493442_m1), Tnnt2 (Mm00441922_m1), and Wnt3 (Mm00437336_m1).

    Techniques: Expressing, Construct, Quantitative RT-PCR

    Tbx6 Expression Promoted Mesoderm and Endoderm Programs and Inhibited Neuroectoderm Genes in Mouse ESCs (A) Hierarchical clustering analysis of iTbx6 T-GFP EBs with or without Dox on days 2 and 4 (n = 2). Dox was added for the first 3 days. (B and C) GO term analyses for the upregulated and downregulated genes in iTbx6 T-GFP EBs on day 2 (B) or 4 (C) of Dox addition. (D) Relative mRNA expression on day 4 in iTbx6 T-GFP EBs with or without Dox addition was determined by qRT-PCR (n = 3, independent experiments). Data were normalized to the values of No Dox. ). (F) ChIP-qPCR analyses in iTbx6 T-GFP EBs with Dox addition on day 4 using antibodies specific for FLAG (Tbx6) or IgG (control) (n = 3). Illustration of the genomic region surrounding Mesp1 . Translated regions are depicted in thick black boxes, untranslated exons are shown in thin black boxes, and introns are shown by black lines. Conserved T-box binding sites are indicated as red lines, and the relative positions of PCR fragments for ChIP-qPCR are represented by black lines. (G) ChIP-qPCR analysis of iTbx6 T-GFP EBs treated with Dox on day 4 using antibodies specific for RNA polymerase II (PolII) or IgG (control) (n = 3). The primers were generated near the transcriptional start site (TSS) of Mesp1 . (H) Time course of mRNA expression for Bmp4 , Nodal , and Wnt3 during the differentiation of ESCs after 12, 24, 48, and 96 hr of Dox treatment, determined by qRT-PCR. Bmp4 was rapidly and strongly upregulated after only 12 hr of Dox administration. (I) Schematic representation of chimeric EB experiments. The iTbx6 T-GFP ESCs (inducer) and T-GFP ESCs (responder) were mixed and cultured for 4 days with Dox addition from days 0–3 in the absence of cytokines before FACS. FACS analyses for Flk1 and PDGFRα expression in day 4 EBs are shown (right).

    Journal: Cell stem cell

    Article Title: Tbx6 Induces Nascent Mesoderm from Pluripotent Stem Cells and Temporally Controls Cardiac versus Somite Lineage Diversification

    doi: 10.1016/j.stem.2018.07.001

    Figure Lengend Snippet: Tbx6 Expression Promoted Mesoderm and Endoderm Programs and Inhibited Neuroectoderm Genes in Mouse ESCs (A) Hierarchical clustering analysis of iTbx6 T-GFP EBs with or without Dox on days 2 and 4 (n = 2). Dox was added for the first 3 days. (B and C) GO term analyses for the upregulated and downregulated genes in iTbx6 T-GFP EBs on day 2 (B) or 4 (C) of Dox addition. (D) Relative mRNA expression on day 4 in iTbx6 T-GFP EBs with or without Dox addition was determined by qRT-PCR (n = 3, independent experiments). Data were normalized to the values of No Dox. ). (F) ChIP-qPCR analyses in iTbx6 T-GFP EBs with Dox addition on day 4 using antibodies specific for FLAG (Tbx6) or IgG (control) (n = 3). Illustration of the genomic region surrounding Mesp1 . Translated regions are depicted in thick black boxes, untranslated exons are shown in thin black boxes, and introns are shown by black lines. Conserved T-box binding sites are indicated as red lines, and the relative positions of PCR fragments for ChIP-qPCR are represented by black lines. (G) ChIP-qPCR analysis of iTbx6 T-GFP EBs treated with Dox on day 4 using antibodies specific for RNA polymerase II (PolII) or IgG (control) (n = 3). The primers were generated near the transcriptional start site (TSS) of Mesp1 . (H) Time course of mRNA expression for Bmp4 , Nodal , and Wnt3 during the differentiation of ESCs after 12, 24, 48, and 96 hr of Dox treatment, determined by qRT-PCR. Bmp4 was rapidly and strongly upregulated after only 12 hr of Dox administration. (I) Schematic representation of chimeric EB experiments. The iTbx6 T-GFP ESCs (inducer) and T-GFP ESCs (responder) were mixed and cultured for 4 days with Dox addition from days 0–3 in the absence of cytokines before FACS. FACS analyses for Flk1 and PDGFRα expression in day 4 EBs are shown (right).

    Article Snippet: Mouse gene expression assays used the following probes: Acta1 (Mm808218_g1), Actn2 (Mm00473657_m1), Bmp4 (Mm00432087_m1), Cdx2 (Mm01212280_m1), Col2a1 (Mm01309565_m1), Eomes (Mm01351985_m1), Flk1 (Mm01222431 m1), Gapdh (Mm99999915_g1), Gsc (Mm00650681_g1), Isl1 (Mm00517585_m1), Lhx1 (Mm01297482_m1), Meox1 (Mm00440285_m1), Mesp1 (Mm00801883_g1), Msgn1 (Mm00490407_s1), Msx1 (Mm00440330_m1), Myf5 (Mm00435125_m1), Myh3 (Mm01332463_m1), Myh6 (Mm00440354_m1), Myh11 (Mm00443013_m1), Myog (Mm00446194_m1), Nkx2.5 (Mm00657783_m1), Nodal ( Mm00443040_m1), Pax3 (Mm00435491_m1), Pax6 (Mm00443081_m1), Pax7 (Mm01354484_m1), Pdgfr a (Mm00440701_m1), Pecam1 (Mm 01242584_m1), Sox1 (Mm00486299_s1), Sox2 (Mm03053810_s1), Sox9 (Mm00448840_m1), Sox17 (Mm00488363_m1), T (Mm00436877_m1), Tbx6 (Mm01278677_m1), Tcf15 (Mm00493442_m1), Tnnt2 (Mm00441922_m1), and Wnt3 (Mm00437336_m1).

    Techniques: Expressing, Quantitative RT-PCR, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Binding Assay, Polymerase Chain Reaction, Generated, Cell Culture, FACS

    Retinoic acid (RA) and BMP4 counteract each other's inhibition of skeletal myogenesis or cardiomyogenesis . P19 cells were mixed with P19[BMP4] or P19[control] cells in the presence of 1% dimethylsulfoxide (DMSO), with or without RA. Panel I : P19[BMP4] cultures treated with RA were fixed on day 9 for immunofluorescence with MF20 antibody (A, B) and counter stained with Hoechst dye (C, D). Magnification is 160x. Panel II : Skeletal myogenesis was quantified for each condition by counting the number of myosin heavy chain +ve bipolar skeletal myocytes, expressed as the percentage of total cells (white bars) and their standard errors ( n = 3). MyoD transcript levels (black bars) were quantified by quantitative polymerase chain reaction and expressed relative to control cultures ( n = 2), * P

    Journal: BMC Biology

    Article Title: Retinoic acid enhances skeletal muscle progenitor formation and bypasses inhibition by bone morphogenetic protein 4 but not dominant negative ?-catenin

    doi: 10.1186/1741-7007-7-67

    Figure Lengend Snippet: Retinoic acid (RA) and BMP4 counteract each other's inhibition of skeletal myogenesis or cardiomyogenesis . P19 cells were mixed with P19[BMP4] or P19[control] cells in the presence of 1% dimethylsulfoxide (DMSO), with or without RA. Panel I : P19[BMP4] cultures treated with RA were fixed on day 9 for immunofluorescence with MF20 antibody (A, B) and counter stained with Hoechst dye (C, D). Magnification is 160x. Panel II : Skeletal myogenesis was quantified for each condition by counting the number of myosin heavy chain +ve bipolar skeletal myocytes, expressed as the percentage of total cells (white bars) and their standard errors ( n = 3). MyoD transcript levels (black bars) were quantified by quantitative polymerase chain reaction and expressed relative to control cultures ( n = 2), * P

    Article Snippet: Plasmid constructs The expression construct phosphoglycerate kinase (PGK)-BMP4 contains a 1.8 Kb EcoRI BMP4 cDNA fragment, containing the complete coding sequence of human BMP4 (Wyeth Pharmaceuticals, MA, USA), driven by the pgk-1 promoter.

    Techniques: Inhibition, Immunofluorescence, Staining, Real-time Polymerase Chain Reaction

    BMP4 inhibits skeletal muscle specification . Panels I and II : P19[BMP4] and P19[control] cells were aggregated in the presence of 0.8% dimethylsulfoxide (DMSO). P19[control] cells were also aggregated in the absence of DMSO to serve as negative controls. On days 0, 6 and 9 (Panel I) and days 1-5 ( Panel II ), total RNA was harvested for northern blot analysis and hybridized with the cDNAs indicated on the right. Panel III : P19 cells were transfected with the TOPFlash reporter, aggregated, and treated with the compounds indicated. Cells were harvested 24 hours later for luciferase assays ( n = 2). Numbers represent the average +/- standard error of mean and statistics were Student's t -test, * P

    Journal: BMC Biology

    Article Title: Retinoic acid enhances skeletal muscle progenitor formation and bypasses inhibition by bone morphogenetic protein 4 but not dominant negative ?-catenin

    doi: 10.1186/1741-7007-7-67

    Figure Lengend Snippet: BMP4 inhibits skeletal muscle specification . Panels I and II : P19[BMP4] and P19[control] cells were aggregated in the presence of 0.8% dimethylsulfoxide (DMSO). P19[control] cells were also aggregated in the absence of DMSO to serve as negative controls. On days 0, 6 and 9 (Panel I) and days 1-5 ( Panel II ), total RNA was harvested for northern blot analysis and hybridized with the cDNAs indicated on the right. Panel III : P19 cells were transfected with the TOPFlash reporter, aggregated, and treated with the compounds indicated. Cells were harvested 24 hours later for luciferase assays ( n = 2). Numbers represent the average +/- standard error of mean and statistics were Student's t -test, * P

    Article Snippet: Plasmid constructs The expression construct phosphoglycerate kinase (PGK)-BMP4 contains a 1.8 Kb EcoRI BMP4 cDNA fragment, containing the complete coding sequence of human BMP4 (Wyeth Pharmaceuticals, MA, USA), driven by the pgk-1 promoter.

    Techniques: Northern Blot, Transfection, Luciferase

    Model of the intersection of retinoic acid (RA), Wnt, and BMP4 signalling during cardiac and skeletal muscle development . BMP4 upregulates Wnt/β-catenin during mesoderm induction (green arrow) and blocks skeletal myogenesis by downregulation of Meox1, Pax3 and myogenic regulatory factor expression (red inhibition arrow). This inhibition can be reversed by RA, which enhances Tob1, Wnt3a, Pax3 and Meox1 expression, activates β-catenin and inhibits BMP4 expression (green arrows). RA receptors bind directly to the Wnt3a, Meox1, and Pax3 regulatory regions (bold green arrows). RA inhibits GATA-4 expression and cardiomyogenesis, likely by inhibiting BMP4 expression and function. Grey arrows indicate previous work [references [ 6 , 8 - 10 , 35 , 38 , 40 , 44 , 102 , 103 ].

    Journal: BMC Biology

    Article Title: Retinoic acid enhances skeletal muscle progenitor formation and bypasses inhibition by bone morphogenetic protein 4 but not dominant negative ?-catenin

    doi: 10.1186/1741-7007-7-67

    Figure Lengend Snippet: Model of the intersection of retinoic acid (RA), Wnt, and BMP4 signalling during cardiac and skeletal muscle development . BMP4 upregulates Wnt/β-catenin during mesoderm induction (green arrow) and blocks skeletal myogenesis by downregulation of Meox1, Pax3 and myogenic regulatory factor expression (red inhibition arrow). This inhibition can be reversed by RA, which enhances Tob1, Wnt3a, Pax3 and Meox1 expression, activates β-catenin and inhibits BMP4 expression (green arrows). RA receptors bind directly to the Wnt3a, Meox1, and Pax3 regulatory regions (bold green arrows). RA inhibits GATA-4 expression and cardiomyogenesis, likely by inhibiting BMP4 expression and function. Grey arrows indicate previous work [references [ 6 , 8 - 10 , 35 , 38 , 40 , 44 , 102 , 103 ].

    Article Snippet: Plasmid constructs The expression construct phosphoglycerate kinase (PGK)-BMP4 contains a 1.8 Kb EcoRI BMP4 cDNA fragment, containing the complete coding sequence of human BMP4 (Wyeth Pharmaceuticals, MA, USA), driven by the pgk-1 promoter.

    Techniques: Expressing, Inhibition

    BMP4 inhibits skeletal but not cardiac myogenesis . Panel I : P19[BMP4] and P19[control] cells were aggregated in the presence of 0.8% dimethylsulfoxide (DMSO). Cells were fixed on day 9, stained with MF20 antibody (A-D), and counter-stained with Hoechst dye to show the nuclei (E-H). Magnification is 400x. Panel II : The number of MHC+ve cells were counted and the average +/- standard error of mean shown. Statistics were Student's t -test, * P

    Journal: BMC Biology

    Article Title: Retinoic acid enhances skeletal muscle progenitor formation and bypasses inhibition by bone morphogenetic protein 4 but not dominant negative ?-catenin

    doi: 10.1186/1741-7007-7-67

    Figure Lengend Snippet: BMP4 inhibits skeletal but not cardiac myogenesis . Panel I : P19[BMP4] and P19[control] cells were aggregated in the presence of 0.8% dimethylsulfoxide (DMSO). Cells were fixed on day 9, stained with MF20 antibody (A-D), and counter-stained with Hoechst dye to show the nuclei (E-H). Magnification is 400x. Panel II : The number of MHC+ve cells were counted and the average +/- standard error of mean shown. Statistics were Student's t -test, * P

    Article Snippet: Plasmid constructs The expression construct phosphoglycerate kinase (PGK)-BMP4 contains a 1.8 Kb EcoRI BMP4 cDNA fragment, containing the complete coding sequence of human BMP4 (Wyeth Pharmaceuticals, MA, USA), driven by the pgk-1 promoter.

    Techniques: Staining

    TWSG1 inhibit up-regulation of hepcidin expression by BMP2/4 in human hepatoma cell line . (A) Relative hepcidin expression as a dosed response to BMP2 (□) and BMP4 (■) are shown. Final BMP concentrations are on the x-axis, and relative

    Journal: Blood

    Article Title: Identification of TWSG1 as a second novel erythroid regulator of hepcidin expression in murine and human cells

    doi: 10.1182/blood-2008-12-195503

    Figure Lengend Snippet: TWSG1 inhibit up-regulation of hepcidin expression by BMP2/4 in human hepatoma cell line . (A) Relative hepcidin expression as a dosed response to BMP2 (□) and BMP4 (■) are shown. Final BMP concentrations are on the x-axis, and relative

    Article Snippet: Hepcidin assays with recombinant proteins were performed under the conditions described by Tanno et al. Human TWSG1 (Peprotech), human bone morphogenic protein 2 (BMP2), and human BMP4 (R & D Systems) were used to determine the effects on hepcidin mRNA expression in human hepatoma cell line (HuH-7) and human primary hepatocytes.

    Techniques: Expressing

    TWSG1 inhibits BMP2/4 up-regulation of hepcidin in primary human hepatocytes . Dose response of relative hepcidin expression levels to (A) BMP2 and (C) BMP4 are shown using primary hepatocytes from 4 separate donors (shaded bars). Final BMP concentrations

    Journal: Blood

    Article Title: Identification of TWSG1 as a second novel erythroid regulator of hepcidin expression in murine and human cells

    doi: 10.1182/blood-2008-12-195503

    Figure Lengend Snippet: TWSG1 inhibits BMP2/4 up-regulation of hepcidin in primary human hepatocytes . Dose response of relative hepcidin expression levels to (A) BMP2 and (C) BMP4 are shown using primary hepatocytes from 4 separate donors (shaded bars). Final BMP concentrations

    Article Snippet: Hepcidin assays with recombinant proteins were performed under the conditions described by Tanno et al. Human TWSG1 (Peprotech), human bone morphogenic protein 2 (BMP2), and human BMP4 (R & D Systems) were used to determine the effects on hepcidin mRNA expression in human hepatoma cell line (HuH-7) and human primary hepatocytes.

    Techniques: Expressing

    Lung epithelial branching morphogenesis was severely impaired in Apc CKO embryos. a Lung epithelial branches at E13.5 were visualized by whole mount E-cadherin immunofluorescence staining. b Altered expression of Fgf10 and Bmp4 in Apc CKO lung at E11.5 was detected by real-time PCR. * P

    Journal: BMC Biology

    Article Title: Mesenchymal adenomatous polyposis coli plays critical and diverse roles in regulating lung development

    doi: 10.1186/s12915-015-0153-1

    Figure Lengend Snippet: Lung epithelial branching morphogenesis was severely impaired in Apc CKO embryos. a Lung epithelial branches at E13.5 were visualized by whole mount E-cadherin immunofluorescence staining. b Altered expression of Fgf10 and Bmp4 in Apc CKO lung at E11.5 was detected by real-time PCR. * P

    Article Snippet: The cultured cells were treated with BMP4 (50 ng/ml to 100 ng/ml final concentration, R & D Systems) overnight and then harvested for RNA and protein analyses.

    Techniques: Immunofluorescence, Staining, Expressing, Real-time Polymerase Chain Reaction

    Twsg1 dose response of hepcidin levels in primary murine hepatocytes . In primary hepatocytes from 4- to 6-week-old C57Bl/6 mice, dosed increased Twsg1 (x-axis) was added, and hepcidin (y-axis; hamp1) mRNA was quantified. * P

    Journal: Blood

    Article Title: Identification of TWSG1 as a second novel erythroid regulator of hepcidin expression in murine and human cells

    doi: 10.1182/blood-2008-12-195503

    Figure Lengend Snippet: Twsg1 dose response of hepcidin levels in primary murine hepatocytes . In primary hepatocytes from 4- to 6-week-old C57Bl/6 mice, dosed increased Twsg1 (x-axis) was added, and hepcidin (y-axis; hamp1) mRNA was quantified. * P

    Article Snippet: Hepcidin assays with recombinant proteins were performed under the conditions described by Tanno et al. Human TWSG1 (Peprotech), human bone morphogenic protein 2 (BMP2), and human BMP4 (R & D Systems) were used to determine the effects on hepcidin mRNA expression in human hepatoma cell line (HuH-7) and human primary hepatocytes.

    Techniques: Mouse Assay

    Model of hepcidin regulation by TWSG1 and GDF15 .

    Journal: Blood

    Article Title: Identification of TWSG1 as a second novel erythroid regulator of hepcidin expression in murine and human cells

    doi: 10.1182/blood-2008-12-195503

    Figure Lengend Snippet: Model of hepcidin regulation by TWSG1 and GDF15 .

    Article Snippet: Hepcidin assays with recombinant proteins were performed under the conditions described by Tanno et al. Human TWSG1 (Peprotech), human bone morphogenic protein 2 (BMP2), and human BMP4 (R & D Systems) were used to determine the effects on hepcidin mRNA expression in human hepatoma cell line (HuH-7) and human primary hepatocytes.

    Techniques:

    TWSG1 inhibit up-regulation of hepcidin expression by BMP2/4 in human hepatoma cell line . (A) Relative hepcidin expression as a dosed response to BMP2 (□) and BMP4 (■) are shown. Final BMP concentrations are on the x-axis, and relative

    Journal: Blood

    Article Title: Identification of TWSG1 as a second novel erythroid regulator of hepcidin expression in murine and human cells

    doi: 10.1182/blood-2008-12-195503

    Figure Lengend Snippet: TWSG1 inhibit up-regulation of hepcidin expression by BMP2/4 in human hepatoma cell line . (A) Relative hepcidin expression as a dosed response to BMP2 (□) and BMP4 (■) are shown. Final BMP concentrations are on the x-axis, and relative

    Article Snippet: Hepcidin assays with recombinant proteins were performed under the conditions described by Tanno et al. Human TWSG1 (Peprotech), human bone morphogenic protein 2 (BMP2), and human BMP4 (R & D Systems) were used to determine the effects on hepcidin mRNA expression in human hepatoma cell line (HuH-7) and human primary hepatocytes.

    Techniques: Expressing

    TWSG1 inhibits BMP2/4 up-regulation of hepcidin in primary human hepatocytes . Dose response of relative hepcidin expression levels to (A) BMP2 and (C) BMP4 are shown using primary hepatocytes from 4 separate donors (shaded bars). Final BMP concentrations

    Journal: Blood

    Article Title: Identification of TWSG1 as a second novel erythroid regulator of hepcidin expression in murine and human cells

    doi: 10.1182/blood-2008-12-195503

    Figure Lengend Snippet: TWSG1 inhibits BMP2/4 up-regulation of hepcidin in primary human hepatocytes . Dose response of relative hepcidin expression levels to (A) BMP2 and (C) BMP4 are shown using primary hepatocytes from 4 separate donors (shaded bars). Final BMP concentrations

    Article Snippet: Hepcidin assays with recombinant proteins were performed under the conditions described by Tanno et al. Human TWSG1 (Peprotech), human bone morphogenic protein 2 (BMP2), and human BMP4 (R & D Systems) were used to determine the effects on hepcidin mRNA expression in human hepatoma cell line (HuH-7) and human primary hepatocytes.

    Techniques: Expressing

    TWSG1 inhibits BMP-mediated Smad phosphorylation . HuH-7 cells were cultured in the presence or absence of 1000 ng/mL TWSG1, 10 ng/mL BMP2, or both for 1 hour. Total cell lysates (20 μg/lane) were immunoblotted using a rabbit antiserum specific

    Journal: Blood

    Article Title: Identification of TWSG1 as a second novel erythroid regulator of hepcidin expression in murine and human cells

    doi: 10.1182/blood-2008-12-195503

    Figure Lengend Snippet: TWSG1 inhibits BMP-mediated Smad phosphorylation . HuH-7 cells were cultured in the presence or absence of 1000 ng/mL TWSG1, 10 ng/mL BMP2, or both for 1 hour. Total cell lysates (20 μg/lane) were immunoblotted using a rabbit antiserum specific

    Article Snippet: Hepcidin assays with recombinant proteins were performed under the conditions described by Tanno et al. Human TWSG1 (Peprotech), human bone morphogenic protein 2 (BMP2), and human BMP4 (R & D Systems) were used to determine the effects on hepcidin mRNA expression in human hepatoma cell line (HuH-7) and human primary hepatocytes.

    Techniques: Cell Culture

    Twsg1 mRNA expression in murine thalassemia . Murine Twsg1 mRNA in (A) spleen, (B) liver, and (C) bone marrow from wild-type mice (WT, n = 7), Hbb th3/+ β-thalassemia intermedia mice ( th3 /+, n = 13), and Hbb th3/th3 β-thalassemia major mice

    Journal: Blood

    Article Title: Identification of TWSG1 as a second novel erythroid regulator of hepcidin expression in murine and human cells

    doi: 10.1182/blood-2008-12-195503

    Figure Lengend Snippet: Twsg1 mRNA expression in murine thalassemia . Murine Twsg1 mRNA in (A) spleen, (B) liver, and (C) bone marrow from wild-type mice (WT, n = 7), Hbb th3/+ β-thalassemia intermedia mice ( th3 /+, n = 13), and Hbb th3/th3 β-thalassemia major mice

    Article Snippet: Hepcidin assays with recombinant proteins were performed under the conditions described by Tanno et al. Human TWSG1 (Peprotech), human bone morphogenic protein 2 (BMP2), and human BMP4 (R & D Systems) were used to determine the effects on hepcidin mRNA expression in human hepatoma cell line (HuH-7) and human primary hepatocytes.

    Techniques: Expressing, Mouse Assay

    TWSG1 expression in human erythropoiesis . (A) Microarray confirmation by quantitative PCR using erythroblasts from 6 separate donors (y-axis, copy number/ng total RNA). Each line associated values from the same donor on separate days. D7, culture day

    Journal: Blood

    Article Title: Identification of TWSG1 as a second novel erythroid regulator of hepcidin expression in murine and human cells

    doi: 10.1182/blood-2008-12-195503

    Figure Lengend Snippet: TWSG1 expression in human erythropoiesis . (A) Microarray confirmation by quantitative PCR using erythroblasts from 6 separate donors (y-axis, copy number/ng total RNA). Each line associated values from the same donor on separate days. D7, culture day

    Article Snippet: Hepcidin assays with recombinant proteins were performed under the conditions described by Tanno et al. Human TWSG1 (Peprotech), human bone morphogenic protein 2 (BMP2), and human BMP4 (R & D Systems) were used to determine the effects on hepcidin mRNA expression in human hepatoma cell line (HuH-7) and human primary hepatocytes.

    Techniques: Expressing, Microarray, Real-time Polymerase Chain Reaction

    Expression of primary dysmenorrhea-related genes by quantitative RT-PCR array on the seventh day before menstruation, and the first and fifth days of menstruation. Compared with the unaffected control group, the primary dysmenorrhea group has the relatively low expression of genes ( BMP6 , GDF5 , GDF11 , NODAL , IL1F5 , IL11 and MSTN ), and high expression of pro-inflammatory cytokines ( IL1B, TNF, IL6, and IL8 ).

    Journal: PLoS ONE

    Article Title: Altered Cytokine Gene Expression in Peripheral Blood Monocytes across the Menstrual Cycle in Primary Dysmenorrhea: A Case-Control Study

    doi: 10.1371/journal.pone.0055200

    Figure Lengend Snippet: Expression of primary dysmenorrhea-related genes by quantitative RT-PCR array on the seventh day before menstruation, and the first and fifth days of menstruation. Compared with the unaffected control group, the primary dysmenorrhea group has the relatively low expression of genes ( BMP6 , GDF5 , GDF11 , NODAL , IL1F5 , IL11 and MSTN ), and high expression of pro-inflammatory cytokines ( IL1B, TNF, IL6, and IL8 ).

    Article Snippet: This mixture was added to 96 wells in an RT2 Profiler™ PCR Array, Human Common Cytokines (PAHS-012A; QIAGEN).

    Techniques: Expressing, Quantitative RT-PCR

    Expression of cardiac markers in somite cells from embryos lacking Myo/Nog cells Somite cultures were prepared from untreated and G8 and complement treated stage 12–13 embryos. Somite cells from untreated embryos formed sarcomeric myosin+, elongated skeletal myotubes (A). Cultures from ablated embryos contained GATA4+ and cardiac troponin I+ (CTpI) cells (B). Some cardiomyocytes contained striations (arrows in B). CTpI+ cells in somite cultures resembled cultured cardiomyocytes from the stage 12 heart (C). In vivo , the somites of G8 ablated (G, H) but not untreated (D, E) stage 12–16 (D, G) and stage 25–26 embryos (E and H) contained GATA4+ and CTpI+ cells. Cardiac troponin T (CTpT) was expressed in stage 25–26 ablated (I) but not untreated embryos (F). Bar, 9 µm.

    Journal: Developmental biology

    Article Title: Myo/Nog Cell Regulation of Bone Morphogenetic Protein Signaling in the Blastocyst is Essential for Normal Morphogenesis and Striated Muscle Lineage Specification

    doi: 10.1016/j.ydbio.2011.08.007

    Figure Lengend Snippet: Expression of cardiac markers in somite cells from embryos lacking Myo/Nog cells Somite cultures were prepared from untreated and G8 and complement treated stage 12–13 embryos. Somite cells from untreated embryos formed sarcomeric myosin+, elongated skeletal myotubes (A). Cultures from ablated embryos contained GATA4+ and cardiac troponin I+ (CTpI) cells (B). Some cardiomyocytes contained striations (arrows in B). CTpI+ cells in somite cultures resembled cultured cardiomyocytes from the stage 12 heart (C). In vivo , the somites of G8 ablated (G, H) but not untreated (D, E) stage 12–16 (D, G) and stage 25–26 embryos (E and H) contained GATA4+ and CTpI+ cells. Cardiac troponin T (CTpT) was expressed in stage 25–26 ablated (I) but not untreated embryos (F). Bar, 9 µm.

    Article Snippet: Double labeling was also carried out with IgG MAbs to Pax3 , sarcomeric myosin heavy chain (MF20 MAb) , neurofilament associated antigen (3A10 MAb) , the notochord marker NOT1, cardiac troponin I (TI-1 MAb) , Shh (5E1 MAb) ( ) (all obtained from the Developmental Studies Hybridoma Bank, Iowa City, IA), MyoD1 (NCL-MyoD1 MAb from Vector Laboratories, Burlingame, CA), BMP2 (Sigma-Aldrich, St. Louis, MO) and cardiac troponin T, isoform Ab-1 (MS-295-PO, Thermo Scientific, Fremont, CA), and the following polyclonal antisera: goat anti-mouse noggin (AF719, R & D Systems, Minneapolis, MN), rabbit anti-mouse chordin (ab24562, Abcam, Cambridge, MA), goat anti-human follistatin (sc-23553, Santa Cruz Biotechnology, Santa Cruz, CA), rabbit anti-human BMP4 (5674R-100, Biovision, Mountain View, CA), rabbit anti-human p-Smad1/5/8 (9511, Cell Signaling Technology, Danvers, MA), rabbit anti-human Wnt3a (ab28472, Abcam), rabbit anti-human GATA4 (ab25992, Abcam) and rabbit anti-human Smad4 (sc-7154, Santa Cruz Biotechnology).

    Techniques: Expressing, Cell Culture, In Vivo

    BMP and Noggin influence neural crest-derived migration and neurite patterning in E11.5 mouse midgut Gut explants were grown in culture with the cut end of the midgut placed onto filter paper to allow outgrowth (see diagram G). Migration and neurite extension from the midgut was examined after 48 hours in culture with GDNF (A, A′, A″), GDNF plus noggin (B, B′, B″), GDNF plus anti-BMP4 blocking antibody (C, C′, C″), GDNF plus BMP2 (D, D′, D″), or GDNF plus BMP4 (E, E′, E″). Immunohistochemistry was performed for TuJ1 (A–E) or Ret (A′–E′). Merged images (A″ – E″). (H) The schematic shows how the extent of migration from the edge of the explant was measured. Seven radiating lines were drawn in standard positions extending from the edge of the explant. The position of the most distant Ret+ cell along these lines was recorded. (F) Mean distance that Ret+ cells migrated from the edge of the explant confirms increased migration with GDNF plus noggin or GDNF plus anti-BMP4 blocking antibody, and reduced migration with GDNF plus BMP4 (n = 12 explants) from 6 separate experiments. Scale bar = 100 μm. (I–L) To confirm the location of Ret+ cells that migrated from the explants, cell nuclei were visualized using Hoechst 33342 dye. Low magnification images of an explant grown in GDNF plus noggin are shown after immunohistochemistry for TuJ1 (I), Ret (J) and Hoechst 33342 (K) with an additional merged image (L). The white box in (L) marks the region shown in (N). (M–P) Show higher magnification images of cells and neurites at the edge of the outgrowth from explants after growth in GDNF (M), GDNF plus noggin (N), GDNF plus BMP4 blocking antibody (O) or GDNF plus BMP4 (P). These images demonstrate cell bodies at the tips of the furthest neurites and highlight BMP4 induced neurite fasciculation (P). * P

    Journal: Developmental biology

    Article Title: BMP signaling regulates murine enteric nervous system precursor migration, neurite fasciculation and patterning via altered Ncam1 polysialic acid addition

    doi: 10.1016/j.ydbio.2006.07.016

    Figure Lengend Snippet: BMP and Noggin influence neural crest-derived migration and neurite patterning in E11.5 mouse midgut Gut explants were grown in culture with the cut end of the midgut placed onto filter paper to allow outgrowth (see diagram G). Migration and neurite extension from the midgut was examined after 48 hours in culture with GDNF (A, A′, A″), GDNF plus noggin (B, B′, B″), GDNF plus anti-BMP4 blocking antibody (C, C′, C″), GDNF plus BMP2 (D, D′, D″), or GDNF plus BMP4 (E, E′, E″). Immunohistochemistry was performed for TuJ1 (A–E) or Ret (A′–E′). Merged images (A″ – E″). (H) The schematic shows how the extent of migration from the edge of the explant was measured. Seven radiating lines were drawn in standard positions extending from the edge of the explant. The position of the most distant Ret+ cell along these lines was recorded. (F) Mean distance that Ret+ cells migrated from the edge of the explant confirms increased migration with GDNF plus noggin or GDNF plus anti-BMP4 blocking antibody, and reduced migration with GDNF plus BMP4 (n = 12 explants) from 6 separate experiments. Scale bar = 100 μm. (I–L) To confirm the location of Ret+ cells that migrated from the explants, cell nuclei were visualized using Hoechst 33342 dye. Low magnification images of an explant grown in GDNF plus noggin are shown after immunohistochemistry for TuJ1 (I), Ret (J) and Hoechst 33342 (K) with an additional merged image (L). The white box in (L) marks the region shown in (N). (M–P) Show higher magnification images of cells and neurites at the edge of the outgrowth from explants after growth in GDNF (M), GDNF plus noggin (N), GDNF plus BMP4 blocking antibody (O) or GDNF plus BMP4 (P). These images demonstrate cell bodies at the tips of the furthest neurites and highlight BMP4 induced neurite fasciculation (P). * P

    Article Snippet: For each gene, qRT-PCR was performed with RNA from E11.5 gut explants that had been maintained in control medium for 48 hours with or without BMP2, BMP4 and noggin as described above (n = 3 under each condition).

    Techniques: Derivative Assay, Migration, Blocking Assay, Immunohistochemistry

    Noggin enhances neural crest migration into the hindgut E11.5 mouse gut explants including the esophagus, stomach, small bowel and colon were cultured for 48 hours (A) under control conditions, or (B) with noggin, (C) anti-BMP4 blocking antibody, (D) BMP2, or (E) BMP4. Whole mount immunohistochemistry for Ret demonstrated NCC within the bowel. White lines indicate the end of the colon. (F) Shows landmarks for measurements: segment “a–b” = the region of colon colonized by NCC; segment “a–c” = the whole length of the hindgut. “a” is the ileo-cecal junction; “b” is the position of the most distal Ret expressing cell at the migration wave front. “c” is the terminal hindgut. Also seen is the extra-intestinal tissue we used to secure the explants to the gel bed. (G) Quantitative analysis demonstrated that noggin or anti-BMP4 blocking antibody treatment increased the percentage of the colon innervated, while BMP treatment reduced the extent of colon innervation. Scale bar = 100 μm. * P

    Journal: Developmental biology

    Article Title: BMP signaling regulates murine enteric nervous system precursor migration, neurite fasciculation and patterning via altered Ncam1 polysialic acid addition

    doi: 10.1016/j.ydbio.2006.07.016

    Figure Lengend Snippet: Noggin enhances neural crest migration into the hindgut E11.5 mouse gut explants including the esophagus, stomach, small bowel and colon were cultured for 48 hours (A) under control conditions, or (B) with noggin, (C) anti-BMP4 blocking antibody, (D) BMP2, or (E) BMP4. Whole mount immunohistochemistry for Ret demonstrated NCC within the bowel. White lines indicate the end of the colon. (F) Shows landmarks for measurements: segment “a–b” = the region of colon colonized by NCC; segment “a–c” = the whole length of the hindgut. “a” is the ileo-cecal junction; “b” is the position of the most distal Ret expressing cell at the migration wave front. “c” is the terminal hindgut. Also seen is the extra-intestinal tissue we used to secure the explants to the gel bed. (G) Quantitative analysis demonstrated that noggin or anti-BMP4 blocking antibody treatment increased the percentage of the colon innervated, while BMP treatment reduced the extent of colon innervation. Scale bar = 100 μm. * P

    Article Snippet: For each gene, qRT-PCR was performed with RNA from E11.5 gut explants that had been maintained in control medium for 48 hours with or without BMP2, BMP4 and noggin as described above (n = 3 under each condition).

    Techniques: Migration, Cell Culture, Blocking Assay, Immunohistochemistry, Expressing

    Expression and distribution of intrathymic BMPs and chordin. (a) Total thymus or isolated stromal cells and thymocytes were subjected to RT-PCR analysis to establish the sources of BMP2, BMP4, BMP7, and chordin in the thymus. GAPDH served as a positive control. H 2 O lanes contained no cDNA. (b–e) Immunoperoxidase staining of normal 3–4-wk-old mouse (C57BL/6) thymus with goat anti-BMP/24 (c and e) or control goat antibody (b and d). BMP2/4 staining (brown) is seen mainly within supcapsular (sc) and medullary (med, outlined in c) regions (original magnification: ×200, b and c) and ‘patchy’ within positive areas (original magnification: ×400, d and e). (f–h) In situ hybridization of normal 3–4-wk-old mouse (C57BL/6) thymus with Tsg sense (f) and antisense (g and h) probes. Tsg expressing cells are distributed throughout the cortex and the medulla. Original magnification: ×100 (f and g), ×200 (h).

    Journal: The Journal of Experimental Medicine

    Article Title: The Developmentally Regulated Expression of Twisted Gastrulation Reveals a Role for Bone Morphogenetic Proteins in the Control of T Cell Development

    doi: 10.1084/jem.20020276

    Figure Lengend Snippet: Expression and distribution of intrathymic BMPs and chordin. (a) Total thymus or isolated stromal cells and thymocytes were subjected to RT-PCR analysis to establish the sources of BMP2, BMP4, BMP7, and chordin in the thymus. GAPDH served as a positive control. H 2 O lanes contained no cDNA. (b–e) Immunoperoxidase staining of normal 3–4-wk-old mouse (C57BL/6) thymus with goat anti-BMP/24 (c and e) or control goat antibody (b and d). BMP2/4 staining (brown) is seen mainly within supcapsular (sc) and medullary (med, outlined in c) regions (original magnification: ×200, b and c) and ‘patchy’ within positive areas (original magnification: ×400, d and e). (f–h) In situ hybridization of normal 3–4-wk-old mouse (C57BL/6) thymus with Tsg sense (f) and antisense (g and h) probes. Tsg expressing cells are distributed throughout the cortex and the medulla. Original magnification: ×100 (f and g), ×200 (h).

    Article Snippet: Thymocyte organ cultures and suspension cultures of mechanically dissociated or trypsinized fetal thymi were set up as hanging drops in inverted Terasaki plates (Nunc) in serum-free AIM-V lymphocyte medium (GIBCO BRL) supplemented with 2 × 10−5 M 2-ME, where indicated in the presence of recombinant BMP2, -4, and -7, chordin, neutralizing anti-BMP4 or BMPR-IA/Fc (all from R & D Systems), TGFβ1 (Sigma-Aldrich), or the CD3ε antibody 2C11 (BD PharMingen).

    Techniques: Expressing, Isolation, Reverse Transcription Polymerase Chain Reaction, Positive Control, Immunoperoxidase Staining, Staining, In Situ Hybridization

    Effects of Myo/Nog cell ablation on the expression of regulators of BMP signaling in the stage 2 embryo Untreated embryos and G8 or D4/complement treated stage X–XII embryos were incubated to stage 2 and double labeled with antibodies. The colors of the fluorescent tags are indicated in each panel. The photograph of the stage 2 embryo (A) illustrates the regions shown in the fluorescence micrographs. Overlap of red and green appears yellow/white in merged images. After ablating G8+ cells in the stage X–XII epiblast, neither G8 (B, C, E–I, K–N), noggin (NogP) (B) nor follistatin (Fol) (C) was detected in the stage 2 embryo. Ablation with the D4 MAb did not affect follistatin expression (D). Staining for chordin (Chor) was reduced along Koller’s sickle (ks) in G8 ablated embryos (E). In untreated stage 2 embryos, BMP4+ cells surrounded Koller’s sickle (F) and BMP2 was present below Koller’s sickle (bks) (G). P-Smad1/5/8 was detected only below Koller’s sickle (H) and did not overlap with G8+ cells in the posterior/medial epiblast (pme) (I). Cytoplasmic staining for Smad4 was present in the pme (J). In G8 ablated embryos, staining for BMP4 was increased in the pme (K), BMP2 remained below Koller’s sickle (L) and p-Smad1/5/8 was expanded into the pme (M) and central epiblast (ce) (N). P-Smad1/5/8 was not present above Koller’s sickle in embryos ablated with the D4 MAb (O). Bar, 1,250 µm in A; 9 µm in B–T.

    Journal: Developmental biology

    Article Title: Myo/Nog Cell Regulation of Bone Morphogenetic Protein Signaling in the Blastocyst is Essential for Normal Morphogenesis and Striated Muscle Lineage Specification

    doi: 10.1016/j.ydbio.2011.08.007

    Figure Lengend Snippet: Effects of Myo/Nog cell ablation on the expression of regulators of BMP signaling in the stage 2 embryo Untreated embryos and G8 or D4/complement treated stage X–XII embryos were incubated to stage 2 and double labeled with antibodies. The colors of the fluorescent tags are indicated in each panel. The photograph of the stage 2 embryo (A) illustrates the regions shown in the fluorescence micrographs. Overlap of red and green appears yellow/white in merged images. After ablating G8+ cells in the stage X–XII epiblast, neither G8 (B, C, E–I, K–N), noggin (NogP) (B) nor follistatin (Fol) (C) was detected in the stage 2 embryo. Ablation with the D4 MAb did not affect follistatin expression (D). Staining for chordin (Chor) was reduced along Koller’s sickle (ks) in G8 ablated embryos (E). In untreated stage 2 embryos, BMP4+ cells surrounded Koller’s sickle (F) and BMP2 was present below Koller’s sickle (bks) (G). P-Smad1/5/8 was detected only below Koller’s sickle (H) and did not overlap with G8+ cells in the posterior/medial epiblast (pme) (I). Cytoplasmic staining for Smad4 was present in the pme (J). In G8 ablated embryos, staining for BMP4 was increased in the pme (K), BMP2 remained below Koller’s sickle (L) and p-Smad1/5/8 was expanded into the pme (M) and central epiblast (ce) (N). P-Smad1/5/8 was not present above Koller’s sickle in embryos ablated with the D4 MAb (O). Bar, 1,250 µm in A; 9 µm in B–T.

    Article Snippet: Double labeling was also carried out with IgG MAbs to Pax3 , sarcomeric myosin heavy chain (MF20 MAb) , neurofilament associated antigen (3A10 MAb) , the notochord marker NOT1, cardiac troponin I (TI-1 MAb) , Shh (5E1 MAb) ( ) (all obtained from the Developmental Studies Hybridoma Bank, Iowa City, IA), MyoD1 (NCL-MyoD1 MAb from Vector Laboratories, Burlingame, CA), BMP2 (Sigma-Aldrich, St. Louis, MO) and cardiac troponin T, isoform Ab-1 (MS-295-PO, Thermo Scientific, Fremont, CA), and the following polyclonal antisera: goat anti-mouse noggin (AF719, R & D Systems, Minneapolis, MN), rabbit anti-mouse chordin (ab24562, Abcam, Cambridge, MA), goat anti-human follistatin (sc-23553, Santa Cruz Biotechnology, Santa Cruz, CA), rabbit anti-human BMP4 (5674R-100, Biovision, Mountain View, CA), rabbit anti-human p-Smad1/5/8 (9511, Cell Signaling Technology, Danvers, MA), rabbit anti-human Wnt3a (ab28472, Abcam), rabbit anti-human GATA4 (ab25992, Abcam) and rabbit anti-human Smad4 (sc-7154, Santa Cruz Biotechnology).

    Techniques: Expressing, Incubation, Labeling, Fluorescence, Staining

    Effects of reintroducing Myo/Nog cells into ablated embryos on regulators of BMP signaling in the stage 2 epiblast G8+ cells isolated from stage X–XII embryos were injected into the epiblast after the indigenous population of G8+ cells was ablated with complement. Embryos were grown to stage 2 and double labeled with antibodies. The photograph of the stage 2 embryo (A) illustrates the regions shown in the fluorescence micrographs. The colors of the fluorescent tags are indicated in each panel. Overlap of red and green appears yellow in merged images. Implanted G8+ cells synthesized noggin in the posterior/medial epiblast (pme) (B) and in the anterior portion of the developing primitive streak (as) (C), restored expression of follistatin in the posterior/lateral epiblast (ple) (D) and inhibited p-Smad1/5/8 accumulation in the epiblast in the pme (E). P-Smad1/5/8 was present below Koller’s sickle (bks) (F). Bar, 900 µm in A; 9 µm in B–F.

    Journal: Developmental biology

    Article Title: Myo/Nog Cell Regulation of Bone Morphogenetic Protein Signaling in the Blastocyst is Essential for Normal Morphogenesis and Striated Muscle Lineage Specification

    doi: 10.1016/j.ydbio.2011.08.007

    Figure Lengend Snippet: Effects of reintroducing Myo/Nog cells into ablated embryos on regulators of BMP signaling in the stage 2 epiblast G8+ cells isolated from stage X–XII embryos were injected into the epiblast after the indigenous population of G8+ cells was ablated with complement. Embryos were grown to stage 2 and double labeled with antibodies. The photograph of the stage 2 embryo (A) illustrates the regions shown in the fluorescence micrographs. The colors of the fluorescent tags are indicated in each panel. Overlap of red and green appears yellow in merged images. Implanted G8+ cells synthesized noggin in the posterior/medial epiblast (pme) (B) and in the anterior portion of the developing primitive streak (as) (C), restored expression of follistatin in the posterior/lateral epiblast (ple) (D) and inhibited p-Smad1/5/8 accumulation in the epiblast in the pme (E). P-Smad1/5/8 was present below Koller’s sickle (bks) (F). Bar, 900 µm in A; 9 µm in B–F.

    Article Snippet: Double labeling was also carried out with IgG MAbs to Pax3 , sarcomeric myosin heavy chain (MF20 MAb) , neurofilament associated antigen (3A10 MAb) , the notochord marker NOT1, cardiac troponin I (TI-1 MAb) , Shh (5E1 MAb) ( ) (all obtained from the Developmental Studies Hybridoma Bank, Iowa City, IA), MyoD1 (NCL-MyoD1 MAb from Vector Laboratories, Burlingame, CA), BMP2 (Sigma-Aldrich, St. Louis, MO) and cardiac troponin T, isoform Ab-1 (MS-295-PO, Thermo Scientific, Fremont, CA), and the following polyclonal antisera: goat anti-mouse noggin (AF719, R & D Systems, Minneapolis, MN), rabbit anti-mouse chordin (ab24562, Abcam, Cambridge, MA), goat anti-human follistatin (sc-23553, Santa Cruz Biotechnology, Santa Cruz, CA), rabbit anti-human BMP4 (5674R-100, Biovision, Mountain View, CA), rabbit anti-human p-Smad1/5/8 (9511, Cell Signaling Technology, Danvers, MA), rabbit anti-human Wnt3a (ab28472, Abcam), rabbit anti-human GATA4 (ab25992, Abcam) and rabbit anti-human Smad4 (sc-7154, Santa Cruz Biotechnology).

    Techniques: Isolation, Injection, Labeling, Fluorescence, Synthesized, Expressing

    Effects of Myo/Nog cell ablation on the expression of regulators of BMP signaling in the stage 2 embryo Untreated embryos and G8 or D4/complement treated stage X–XII embryos were incubated to stage 2 and double labeled with antibodies. The colors of the fluorescent tags are indicated in each panel. The photograph of the stage 2 embryo (A) illustrates the regions shown in the fluorescence micrographs. Overlap of red and green appears yellow/white in merged images. After ablating G8+ cells in the stage X–XII epiblast, neither G8 (B, C, E–I, K–N), noggin (NogP) (B) nor follistatin (Fol) (C) was detected in the stage 2 embryo. Ablation with the D4 MAb did not affect follistatin expression (D). Staining for chordin (Chor) was reduced along Koller’s sickle (ks) in G8 ablated embryos (E). In untreated stage 2 embryos, BMP4+ cells surrounded Koller’s sickle (F) and BMP2 was present below Koller’s sickle (bks) (G). P-Smad1/5/8 was detected only below Koller’s sickle (H) and did not overlap with G8+ cells in the posterior/medial epiblast (pme) (I). Cytoplasmic staining for Smad4 was present in the pme (J). In G8 ablated embryos, staining for BMP4 was increased in the pme (K), BMP2 remained below Koller’s sickle (L) and p-Smad1/5/8 was expanded into the pme (M) and central epiblast (ce) (N). P-Smad1/5/8 was not present above Koller’s sickle in embryos ablated with the D4 MAb (O). Bar, 1,250 µm in A; 9 µm in B–T.

    Journal: Developmental biology

    Article Title: Myo/Nog Cell Regulation of Bone Morphogenetic Protein Signaling in the Blastocyst is Essential for Normal Morphogenesis and Striated Muscle Lineage Specification

    doi: 10.1016/j.ydbio.2011.08.007

    Figure Lengend Snippet: Effects of Myo/Nog cell ablation on the expression of regulators of BMP signaling in the stage 2 embryo Untreated embryos and G8 or D4/complement treated stage X–XII embryos were incubated to stage 2 and double labeled with antibodies. The colors of the fluorescent tags are indicated in each panel. The photograph of the stage 2 embryo (A) illustrates the regions shown in the fluorescence micrographs. Overlap of red and green appears yellow/white in merged images. After ablating G8+ cells in the stage X–XII epiblast, neither G8 (B, C, E–I, K–N), noggin (NogP) (B) nor follistatin (Fol) (C) was detected in the stage 2 embryo. Ablation with the D4 MAb did not affect follistatin expression (D). Staining for chordin (Chor) was reduced along Koller’s sickle (ks) in G8 ablated embryos (E). In untreated stage 2 embryos, BMP4+ cells surrounded Koller’s sickle (F) and BMP2 was present below Koller’s sickle (bks) (G). P-Smad1/5/8 was detected only below Koller’s sickle (H) and did not overlap with G8+ cells in the posterior/medial epiblast (pme) (I). Cytoplasmic staining for Smad4 was present in the pme (J). In G8 ablated embryos, staining for BMP4 was increased in the pme (K), BMP2 remained below Koller’s sickle (L) and p-Smad1/5/8 was expanded into the pme (M) and central epiblast (ce) (N). P-Smad1/5/8 was not present above Koller’s sickle in embryos ablated with the D4 MAb (O). Bar, 1,250 µm in A; 9 µm in B–T.

    Article Snippet: Double labeling was also carried out with IgG MAbs to Pax3 , sarcomeric myosin heavy chain (MF20 MAb) , neurofilament associated antigen (3A10 MAb) , the notochord marker NOT1, cardiac troponin I (TI-1 MAb) , Shh (5E1 MAb) ( ) (all obtained from the Developmental Studies Hybridoma Bank, Iowa City, IA), MyoD1 (NCL-MyoD1 MAb from Vector Laboratories, Burlingame, CA), BMP2 (Sigma-Aldrich, St. Louis, MO) and cardiac troponin T, isoform Ab-1 (MS-295-PO, Thermo Scientific, Fremont, CA), and the following polyclonal antisera: goat anti-mouse noggin (AF719, R & D Systems, Minneapolis, MN), rabbit anti-mouse chordin (ab24562, Abcam, Cambridge, MA), goat anti-human follistatin (sc-23553, Santa Cruz Biotechnology, Santa Cruz, CA), rabbit anti-human BMP4 (5674R-100, Biovision, Mountain View, CA), rabbit anti-human p-Smad1/5/8 (9511, Cell Signaling Technology, Danvers, MA), rabbit anti-human Wnt3a (ab28472, Abcam), rabbit anti-human GATA4 (ab25992, Abcam) and rabbit anti-human Smad4 (sc-7154, Santa Cruz Biotechnology).

    Techniques: Expressing, Incubation, Labeling, Fluorescence, Staining

    Effects of Myo/Nog cell ablation on the expression of regulators of BMP signaling in gastrulating embryos Untreated embryos (untx) and embryos incubated with complement (cp) only or D4 and complement at stage X–XII displayed a normal primitive streak (ps) by stage 3–4 (A–C). G8 ablated embryos had primitive streaks of abnormal morphology and/or positioning (D, E). Stage 3–4 embryos were double labeled with antibodies. The photographs of the embryos in A and D demonstrate the regions shown in the photomicrographs of F–O and P–T, respectively. The colors of the fluorescent tags are indicated in each panel. Overlap of red and green appears yellow in merged images. In untreated embryos, G8+/noggin+ (NogP) cells were present in the primitive streak (ps) (F) and Hensen’s node (hn) (G). Chordin (Chor) was found in the primitive streak (H) and Hensen’s node (I). Staining for follistatin (Fol) was observed in the anterior primitive streak (J) and above Hensen’s node (ahn) (K). BMP4 (L) and p-Smad1/5/8 (N) were absent in Hensen’s node but present in the lateral ectoderm (ec) (M and O). Staining for chordin, follistatin, BMP4 and p-Smad1/5/8 did not overlap with G8 (H–O). Neither G8, noggin nor follistatin was detected in Hensen’s node in embryos ablated with G8 and complement (P and Q). Staining for chordin was weaker in Hensen’s node of G8 ablated embryos (R) compared to untreated embryos (I). BMP4 (S) and p-Smad1/5/8 (T) were present above Hensen’s node in G8 ablated embryos. Bar, 135 µm in A–E and 9 µm in F–T.

    Journal: Developmental biology

    Article Title: Myo/Nog Cell Regulation of Bone Morphogenetic Protein Signaling in the Blastocyst is Essential for Normal Morphogenesis and Striated Muscle Lineage Specification

    doi: 10.1016/j.ydbio.2011.08.007

    Figure Lengend Snippet: Effects of Myo/Nog cell ablation on the expression of regulators of BMP signaling in gastrulating embryos Untreated embryos (untx) and embryos incubated with complement (cp) only or D4 and complement at stage X–XII displayed a normal primitive streak (ps) by stage 3–4 (A–C). G8 ablated embryos had primitive streaks of abnormal morphology and/or positioning (D, E). Stage 3–4 embryos were double labeled with antibodies. The photographs of the embryos in A and D demonstrate the regions shown in the photomicrographs of F–O and P–T, respectively. The colors of the fluorescent tags are indicated in each panel. Overlap of red and green appears yellow in merged images. In untreated embryos, G8+/noggin+ (NogP) cells were present in the primitive streak (ps) (F) and Hensen’s node (hn) (G). Chordin (Chor) was found in the primitive streak (H) and Hensen’s node (I). Staining for follistatin (Fol) was observed in the anterior primitive streak (J) and above Hensen’s node (ahn) (K). BMP4 (L) and p-Smad1/5/8 (N) were absent in Hensen’s node but present in the lateral ectoderm (ec) (M and O). Staining for chordin, follistatin, BMP4 and p-Smad1/5/8 did not overlap with G8 (H–O). Neither G8, noggin nor follistatin was detected in Hensen’s node in embryos ablated with G8 and complement (P and Q). Staining for chordin was weaker in Hensen’s node of G8 ablated embryos (R) compared to untreated embryos (I). BMP4 (S) and p-Smad1/5/8 (T) were present above Hensen’s node in G8 ablated embryos. Bar, 135 µm in A–E and 9 µm in F–T.

    Article Snippet: Double labeling was also carried out with IgG MAbs to Pax3 , sarcomeric myosin heavy chain (MF20 MAb) , neurofilament associated antigen (3A10 MAb) , the notochord marker NOT1, cardiac troponin I (TI-1 MAb) , Shh (5E1 MAb) ( ) (all obtained from the Developmental Studies Hybridoma Bank, Iowa City, IA), MyoD1 (NCL-MyoD1 MAb from Vector Laboratories, Burlingame, CA), BMP2 (Sigma-Aldrich, St. Louis, MO) and cardiac troponin T, isoform Ab-1 (MS-295-PO, Thermo Scientific, Fremont, CA), and the following polyclonal antisera: goat anti-mouse noggin (AF719, R & D Systems, Minneapolis, MN), rabbit anti-mouse chordin (ab24562, Abcam, Cambridge, MA), goat anti-human follistatin (sc-23553, Santa Cruz Biotechnology, Santa Cruz, CA), rabbit anti-human BMP4 (5674R-100, Biovision, Mountain View, CA), rabbit anti-human p-Smad1/5/8 (9511, Cell Signaling Technology, Danvers, MA), rabbit anti-human Wnt3a (ab28472, Abcam), rabbit anti-human GATA4 (ab25992, Abcam) and rabbit anti-human Smad4 (sc-7154, Santa Cruz Biotechnology).

    Techniques: Expressing, Incubation, Labeling, Staining

    Effects of Myo/Nog cell ablation on the expression of markers for skeletal muscle, neurons and notochord cells Stage X–XII embryos were treated with complement (cp) to ablate G8+ or D4+ cells, grown to stage 25–26, sectioned and labeled with antibodies. The colors of the fluorescent tags are indicated in each panel. The areas illustrated in the low magnification images of H E stained sections are shown in the fluorescence micrographs. Overlap of red and green appears yellow in merged images. D4 ablated embryos contained G8+/noggin+ (Nog) cells (C) and cells with follistatin (Fol) (E), Pax3 (G), MyoD (I) and sarcomeric myosin (K) in the dermomyotome/myotome (dmm). Embryos ablated with the G8 MAb lacked G8 (D), noggin (D), follistatin (F), Pax3 (H), MyoD (J) and sarcomeric myosin (L) in the somites. Staining for neurofilament associated antigen (nfa) in the neural tube (N) and NOT1 in the notochord (P) were reduced in G8 ablated embryos compared to D4 ablated embryos (M and O). Bar, 27 µm in A; 56 µm in B; 9 µm in C–P.

    Journal: Developmental biology

    Article Title: Myo/Nog Cell Regulation of Bone Morphogenetic Protein Signaling in the Blastocyst is Essential for Normal Morphogenesis and Striated Muscle Lineage Specification

    doi: 10.1016/j.ydbio.2011.08.007

    Figure Lengend Snippet: Effects of Myo/Nog cell ablation on the expression of markers for skeletal muscle, neurons and notochord cells Stage X–XII embryos were treated with complement (cp) to ablate G8+ or D4+ cells, grown to stage 25–26, sectioned and labeled with antibodies. The colors of the fluorescent tags are indicated in each panel. The areas illustrated in the low magnification images of H E stained sections are shown in the fluorescence micrographs. Overlap of red and green appears yellow in merged images. D4 ablated embryos contained G8+/noggin+ (Nog) cells (C) and cells with follistatin (Fol) (E), Pax3 (G), MyoD (I) and sarcomeric myosin (K) in the dermomyotome/myotome (dmm). Embryos ablated with the G8 MAb lacked G8 (D), noggin (D), follistatin (F), Pax3 (H), MyoD (J) and sarcomeric myosin (L) in the somites. Staining for neurofilament associated antigen (nfa) in the neural tube (N) and NOT1 in the notochord (P) were reduced in G8 ablated embryos compared to D4 ablated embryos (M and O). Bar, 27 µm in A; 56 µm in B; 9 µm in C–P.

    Article Snippet: Double labeling was also carried out with IgG MAbs to Pax3 , sarcomeric myosin heavy chain (MF20 MAb) , neurofilament associated antigen (3A10 MAb) , the notochord marker NOT1, cardiac troponin I (TI-1 MAb) , Shh (5E1 MAb) ( ) (all obtained from the Developmental Studies Hybridoma Bank, Iowa City, IA), MyoD1 (NCL-MyoD1 MAb from Vector Laboratories, Burlingame, CA), BMP2 (Sigma-Aldrich, St. Louis, MO) and cardiac troponin T, isoform Ab-1 (MS-295-PO, Thermo Scientific, Fremont, CA), and the following polyclonal antisera: goat anti-mouse noggin (AF719, R & D Systems, Minneapolis, MN), rabbit anti-mouse chordin (ab24562, Abcam, Cambridge, MA), goat anti-human follistatin (sc-23553, Santa Cruz Biotechnology, Santa Cruz, CA), rabbit anti-human BMP4 (5674R-100, Biovision, Mountain View, CA), rabbit anti-human p-Smad1/5/8 (9511, Cell Signaling Technology, Danvers, MA), rabbit anti-human Wnt3a (ab28472, Abcam), rabbit anti-human GATA4 (ab25992, Abcam) and rabbit anti-human Smad4 (sc-7154, Santa Cruz Biotechnology).

    Techniques: Expressing, Labeling, Staining, Fluorescence

    Effect of Chordin-like 1 and SB203580 on expression of p21 Cip1/WAF1 , p53, phospho-Rb, and Apo J. A , Western blot analysis of the protein levels of p21 Cip1/WAF1 , p53, and phospho-Rb ( pRb ) in oxidant-treated ARPE-19 cells, with or without Chordin-like

    Journal: The Journal of Biological Chemistry

    Article Title: BMP4 Mediates Oxidative Stress-induced Retinal Pigment Epithelial Cell Senescence and Is Overexpressed in Age-related Macular Degeneration

    doi: 10.1074/jbc.M809393200

    Figure Lengend Snippet: Effect of Chordin-like 1 and SB203580 on expression of p21 Cip1/WAF1 , p53, phospho-Rb, and Apo J. A , Western blot analysis of the protein levels of p21 Cip1/WAF1 , p53, and phospho-Rb ( pRb ) in oxidant-treated ARPE-19 cells, with or without Chordin-like

    Article Snippet: Blocking or Inhibiting Treatment —ARPE-19 cells were treated with 0.1 μg/ml recombinant Chordin-like 1 protein (R & D Systems) or treated with 10 μ m SB203580 (EMD, Gibbstown, NJ) during and after stressor treatments in ARPE medium with 10% fetal bovine serum.

    Techniques: Expressing, Western Blot

    Cell cycle analysis of ARPE-19 cells after oxidant treatment and oxidant plus inhibitors. ARPE-19 cells treated with oxidants alone or co-treated with oxidants and BMP4 antagonist Chordin-like 1 ( CHL1 ) or phospho-p38 inhibitor SB203580 ( SB ) were stained

    Journal: The Journal of Biological Chemistry

    Article Title: BMP4 Mediates Oxidative Stress-induced Retinal Pigment Epithelial Cell Senescence and Is Overexpressed in Age-related Macular Degeneration

    doi: 10.1074/jbc.M809393200

    Figure Lengend Snippet: Cell cycle analysis of ARPE-19 cells after oxidant treatment and oxidant plus inhibitors. ARPE-19 cells treated with oxidants alone or co-treated with oxidants and BMP4 antagonist Chordin-like 1 ( CHL1 ) or phospho-p38 inhibitor SB203580 ( SB ) were stained

    Article Snippet: Blocking or Inhibiting Treatment —ARPE-19 cells were treated with 0.1 μg/ml recombinant Chordin-like 1 protein (R & D Systems) or treated with 10 μ m SB203580 (EMD, Gibbstown, NJ) during and after stressor treatments in ARPE medium with 10% fetal bovine serum.

    Techniques: Cell Cycle Assay, Staining

    Examples of cell type independent, human ES cell and B cell specific Myc targets. ( A ) Gene expression levels (at log2 scale) of FBL , LIN28 and BLMH in the absence and presence of Myc induction in P493-6 B cells (measured by Affymetrix Exon arrays and Affymetrix U133 Plus 2.0 arrays), and log2 gene expression fold changes between hESC and trophoblasts (measured by Agilent microarrays) are shown. Error bars correspond to standard deviations of replicate samples. ( B ) ChIP-chip binding signals in human P493-6 B cells and H9 ES cells, and ENCODE ChIP-seq binding signals in three human cancer cell lines. For ChIP-chip, TileMap moving average statistic [62] m was computed for each probe using normalized log2 probe intensities, and 2 m was displayed as the intensity measure. For ChIP-seq, a 100 bp sliding window was used to scan the genome. Read count in each window was shown at a 25 bp step size. E-box motifs CACGTG (black) and CANNTG (red) were mapped to peak regions and are also shown. ( C ) ChIP-chip [34] and ChIP-seq [40] binding signals in mouse ES cells.

    Journal: PLoS ONE

    Article Title: Cell-Type Independent MYC Target Genes Reveal a Primordial Signature Involved in Biomass Accumulation

    doi: 10.1371/journal.pone.0026057

    Figure Lengend Snippet: Examples of cell type independent, human ES cell and B cell specific Myc targets. ( A ) Gene expression levels (at log2 scale) of FBL , LIN28 and BLMH in the absence and presence of Myc induction in P493-6 B cells (measured by Affymetrix Exon arrays and Affymetrix U133 Plus 2.0 arrays), and log2 gene expression fold changes between hESC and trophoblasts (measured by Agilent microarrays) are shown. Error bars correspond to standard deviations of replicate samples. ( B ) ChIP-chip binding signals in human P493-6 B cells and H9 ES cells, and ENCODE ChIP-seq binding signals in three human cancer cell lines. For ChIP-chip, TileMap moving average statistic [62] m was computed for each probe using normalized log2 probe intensities, and 2 m was displayed as the intensity measure. For ChIP-seq, a 100 bp sliding window was used to scan the genome. Read count in each window was shown at a 25 bp step size. E-box motifs CACGTG (black) and CANNTG (red) were mapped to peak regions and are also shown. ( C ) ChIP-chip [34] and ChIP-seq [40] binding signals in mouse ES cells.

    Article Snippet: RNA samples were submitted to the Johns Hopkins Microarray Core Facility to be hybridized to Agilent Whole Human Genome 44K microarrays (H9 cells) or Affymetrix Human Genome U133 Plus 2.0 or Human Exon 1.0 ST Arrays (P493-6 cells).

    Techniques: Expressing, Chromatin Immunoprecipitation, Binding Assay