Anti-GAL4抗体- ChIP Grade (ab1396)

概述

  • 产品名称Anti-GAL4抗体- ChIP Grade
    参阅全部 GAL4 一抗
  • 描述
    兔多克隆抗体to GAL4 - ChIP Grade
  • 特异性Customers feedbacks suggests that this antibody would not provide satisfactory results in Drosophila melanogaster.
  • 经测试应用适用于: WB, ChIPmore details
  • 种属反应性
    与反应: Saccharomyces cerevisiae
    不与反应: Drosophila melanogaster
  • 免疫原

    Synthetic peptide corresponding to Saccharomyces cerevisiae GAL4 aa 100-200 conjugated to keyhole limpet haemocyanin.
    (Peptide available as ab23612)

  • 阳性对照
    • This antibody gave a positive signal with GAL4-VP16 recombinant protein.

应用

Our Abpromise guarantee covers the use of ab1396 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

应用 Ab评论 说明
WB Use a concentration of 1 µg/ml. Predicted molecular weight: 99 kDa.
ChIP Use at an assay dependent dilution.

靶标

  • 相关性Function: This protein is a positive regulator for the gene expression of the galactose-induced genes such as GAL1, GAL2, GAL7, GAL10, and MEL1 which encode for the enzymes used to convert galactose to glucose. It recognizes a 17 base pair sequence in (5'-CGGRNNRCYNYNCNCCG-3') the upstream activating sequence (UAS-G) of these genes. Subunit structure: Binds DNA as a homodimer. Interacts directly with the mediator subunits GAL11/MED15 and SRB4/MED17. Domain: The 9aaTAD motif (residues 862 to 870) is a transactivation domain present in a large number of yeast and animal transcription factors. Post-translational modification: Association between GAL11 and GAL4 may serve to expedite phosphorylation of GAL4.
  • 细胞定位Nuclear
  • 数据库链接
    • 别名
      • Gal4p antibody
      • GAL81 antibody
      • Regulatory protein GAL4 antibody

    Anti-GAL4 antibody - ChIP Grade 图像



    • Predicted band size : 99 kDa
      Developed using the ECL technique. Performed under reducing conditions with exposure time of 5 mins. The samples run on a 4-20% gradient gel. All blocking and antibody incubation steps done in 5% milk in 20mM Tris-HCl plus 0.1% TWEEN-20.
      Sample 1: Marker. Sample 2A: Whole cell yeast lysate control from untransformed SEY6210 pep4-3 strain (a kind gift from Prof. Tom Stevens from University of Oregon). Sample 2B: Whole cell yeast lysate from SEY6210 pep4-3 strain transformed with pJK22-pGBDU plasmid containing GAL4 DNA Binding domain fused to the VPS60 Gene (a kind gift from Prof. Tom Stevens from University of Oregon). Sample 2C: Whole cell yeast lysate from SEY6210 pep4-3 strain transformed with pJk23-pGAD plasmid containing GAL4 Active Domain fused to the VPS60 Gene (a kind gift from Prof. Tom Stevens from University of Oregon).
      Primary: Lane 1: none. Lane 2: Anti-GAL4BD (ab1396) antibody at 1ug/ml. Secondary: HRP conjugated Goat anti-Rabbit secondary antibody at 1/10000 dilution. Predicted band size: 99 kDa. Additional bands: 56 kDa (possible isoform, Fusion Protein)
    • A stably transfected 293T human cell line harbouring the GAL4 upstream activation sequence was transiently transfected with a V5 or T7- tagged GAL4 DNA Binding Domain construct. 48 hours post transfection Chromatin was prepared according to the Abcam X-ChIP protocol. The ChIP was performed with 25 ug chromatin, 5ug of antibody and 20 ul of Protein A/G beads. A non-specific antibody was used as the negative control. The immunoprecipitated DNA was quantified by real time PCR (SYBR Green approach).
    • S. cerevisiae cells were incubated in raffinose-containing media then transferred to galactose-containing media to activate transcription of galactose activated genes. IP was performed with 500µl of cell extract incubated overnight with 5µl of ab1396 at 4°C, followed by the addition of 25µl of protein A sepharose beads and incubation for 2 hours (room temp). The subsequently purified DNA was analysed with 3 pairs of primers (see diagram). The results show that Gal4 binds specifically to the UAS region of Gal1-Gal10 in both raffinose and galactose, despite only 500-600bp between the UAS and 5' primer.

    • Anti-GAL4 antibody - ChIP Grade (ab1396) at 1 µg/ml + GAL4-VP16 Recombinant Protein at 0.1 µg

      Secondary
      Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution

      Performed under reducing conditions.

      Predicted band size : 99 kDa
      Observed band size : 32 kDa (why is the actual band size different from the predicted?)


      Exposure time : 2 minutes
      ab1396 was tested against GAL4-VP16 Recombinant Protein predicted to run at 28 kDa.

    Anti-GAL4 antibody - ChIP Grade (ab1396)参考文献

    This product has been referenced in:
    • Chen X  et al. Activation of a human chromosomal replication origin by protein tethering. Nucleic Acids Res 41:6460-74 (2013). WB . Read more (PubMed: 23658226) »
    • Inturi R  et al. Adenovirus precursor pVII protein stability is regulated by its propeptide sequence. PLoS One 8:e80617 (2013). WB . Read more (PubMed: 24260437) »

    See all 5 Publications for this product

    Product Wall

    Abcam has not validated the combination of species/application used in this Abreview.
    Application IHC - Wholemount
    Sample Fruit fly (Drosophila melanogaster) Tissue (Wholemount brain tissue)
    Specification Wholemount brain tissue
    Username

    Abcam user community

    Verified customer

    提交于 Jan 06 2014

    Thank you for contacting us.
    We have 2 Gal4 products available in catalogue. One is ab23612 which was used to produce ab1396 and other is ab81879 which is a fusion protein. These products can be used a positive control.
    I hope this information...

    Read More
    Abcam has not validated the combination of species/application used in this Abreview.
    Application Immunohistochemistry (Frozen sections)
    Sample Fruit fly (Drosophila melanogaster) Tissue sections (eye imaginal disc)
    Specification eye imaginal disc
    Fixative Paraformaldehyde
    Permeabilization Yes - tritonX
    Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: ~20°C
    Username

    Ms. Vilaiwan Fernandes

    Verified customer

    提交于 Nov 16 2011

    Thanks for your email. Yes, I do think you're right. I removed Drosophila from the datasheets. If you have any other questions or concerns, just let me know.

    Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"