Product nameAnti-GAL4 antibody - ChIP Grade
See all GAL4 primary antibodies
DescriptionRabbit polyclonal to GAL4 - ChIP Grade
SpecificityCustomers feedbacks suggests that this antibody would not provide satisfactory results in Drosophila melanogaster.
Tested applicationsSuitable for: WB, ChIPmore details
Species reactivityReacts with: Saccharomyces cerevisiae
Does not react with: Drosophila melanogaster
Synthetic peptide corresponding to Saccharomyces cerevisiae GAL4 aa 100-200 conjugated to keyhole limpet haemocyanin.
(Peptide available as
- This antibody gave a positive signal with GAL4-VP16 recombinant protein.
Storage instructionsShipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
Storage bufferPreservative: 0.02% Sodium Azide
Constituents: 1% BSA, PBS. pH 7.4
Concentration information loading...
PurityImmunogen affinity purified
- Epigenetics and Nuclear Signaling
- Polymerase associated factors
- Pol II Transcription
- Pathways and Processes
- Metabolic signaling pathways
- Energy transfer pathways
- Energy Metabolism
ChIP Related Products
Immunizing Peptide (Blocking)
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.
|WB||Use a concentration of 1 µg/ml. Predicted molecular weight: 99 kDa.|
|ChIP||Use at an assay dependent dilution.|
RelevanceFunction: 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.
- Gal4p antibody
- GAL81 antibody
- Regulatory protein GAL4 antibody
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
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.
This product has been referenced in:
- Li J et al. Development of a membrane-anchored ligand and receptor yeast two-hybrid system for ligand-receptor interaction identification. Sci Rep 6:35631 (2016). WB . Read more (PubMed: 27762338) »
- Roedgaard M et al. DNA Topoisomerases Are Required for Preinitiation Complex Assembly during GAL Gene Activation. PLoS One 10:e0132739 (2015). WB, ChIP ; Saccharomyces cerevisiae . Read more (PubMed: 26173127) »