RAD9 Homolog A (S. Pombe) (RAD9A) (S1129) Antikörper

Antigen: RAD9 Homolog A (S. Pombe) (RAD9A)

Synonyme: RAD9, SMC1, SMCB, CDLS2, SB1.8, SMC1L1, DXS423E, KIAA0178, MGC138332, SMC1alpha, DKFZp686L19178, RAD9A, Rad9a, rad9a, MGC89062

Epitope: S1129

Klonalität: Polyklonal

Wirt: Kaninchen

Reaktivität: Hefe (Saccharomyces Cerevisiae)

Applikation: ELISA, Western Blot (WB), Immunpräzipitation (IP)

Produktnummer: ABIN106406

Produktbeschreibung

Weitere Bezeichnung: Rad9 S1129

Immunogen: This affinity purified antibody was prepared from whole rabbit serum produced by repeated immunizations with a synthetic peptide corresponding to aa 1125-1139 of 1309 of yeast Rad9 protein conjugated to KLH.

Format: Liquid (sterile filtered)

Beschreibung: Rad9 is required for the MEC1/TEL1-dependent activation of Saccharomyces cerevisiae DNA damage checkpoint pathways mediated by Rad53 and Chk1. DNA damage induces Rad9 phosphorylation, and Rad53 specifically associates with phosphorylated Rad9. Cells have evolved multiple strategies for tolerating genomic damage. The most important of these are numerous repair systems that remove or bypass potentially mutagenic DNA lesions. Another cellular strategy is to delay cell-cycle transitions at multiple points. The genetic control of these delays, termed `checkpoints', was first established in budding yeast where it was shown that the RAD9 gene functions in G 2 /M arrest after irradiation with X-rays. Subsequently, it has become clear that Rad9 also functions at the G 1 /S, intra-S and mid-anaphase checkpoints. Defects in checkpoint regulation can lead to genome instability and, in higher eukaryotes, neoplastic transformation. Rad9 also controls the transcriptional induction of a DNA damage regulon (DDR). Rad9 may also have a pro-apoptotic function. This is suggested in that Rad9 from Schizosaccharomyces pombe (SpRad9) contains a group of amino acids with similarity to the Bcl-2 homology 3 death domain, which is required for SpRad9 interaction with human Bcl-2 and apoptosis induction in human cells. Overexpression of Bcl-2 in S. pombe inhibits cell growth independently of rad9, but enhances resistance of rad9-null cells to methyl methanesulfonate, ultraviolet and ionizing radiation. Mature yeast Rad9 is reported to have an apparent molecular weight of ~148kDa. The human homolog is reported at 48.5 kDa.

Spezifität: This affinity purified antibody is directed against an internal sequence of yeast Rad9 at the S1129 residue. The product was affinity purified from monospecific antiserum by immunoaffinity purification. Antiserum was purified against the immunizing peptide. This pan reactive polyclonal antibody reacts equally with both phosphorylated and non- phosphorylated yeast Rad9 at S1129. Cross reactivity with Rad9 homologs from other sources has not been determined.

Anwendungen

Applikationshinweise: Recommended Dilutions: This product has been assayed against 0.1 ug of immunizing peptide (S1129) in a standard capture ELISA using TMB (3,3',5,5'-Tetramethylbenizidine) code TMBE-100 as a substrate for 30 minutes at room temperature. A working dilution of 1:5,000 is suggested for this product. Reactivity was detected against both the phosphorylated and non-phosphorylated form (S1129 and pS1129) of the immunizing peptide. This antibody appears to be pan reactive for both forms of the protein. Dilute the antibody 1:100 to 1:500 for immunoblotting. Researchers should determine optimal titers for other applications. Note: This pan reactive polyclonal antibody was tested by immunoblotting and ELISA. Data from both immunoblotting and ELISA indicate the antibody is pan reactive with both the phosphorylated and non-phosphorylated forms of the peptide and protein. Immunoblotting detects yeast Rad9 protein with unknown reactivity to human and mouse homologs. Although not tested, this antibody is likely functional by immunohistochemistry and immunoprecipitation.

Konzentration: 0.74 mg/ml (by UV absorbance at 280 nm)

Buffer: Stabilizer: None. Preservative: 0.01% (w/v) Sodium Azide. Buffer: 0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2.

Lagerung: Store vial at -20° C prior to opening. Centrifuge product if not completely clear after standing at room temperature. Dilute only prior to immediate use. For extended storage aliquot contents and freeze at -20° C or below. Avoid cycles of freezing and thawing. Expiration date is one (1) year from date of opening.

Beschränkungen: Nur für Forschungszwecke einsetzbar

Bilder

Immunoblotting. Affinity purified pan reactive antibody to yeast Rad9 at S1129 was used at a 1:200 dilution incubated 8 h at room temperature to detect Rad9 by Western blot. Lanes were loaded with 50 ng each of recombinant GST fusion protein containing S. cerevisiae Rad9 (aa 991-1309 ~60 kDa) on a 4-20% Criterion gel for SDS-PAGE as follows: Lane 1 - non-phosphorylated wild type yeast Rad9, Lane 2 - in vitro phosphorylated wild type yeast Rad9, Lane 3 - non-phosphorylated S1129A/S1260A double mutant Rad9, Lane 4 - in vitro phosphorylated S1129A/S1260A double mutant. Phosphorylation of Rad9 was by treatment with ATP and Rad53 kinase. Detection occurred using a 1:5,000 dilution of IRDye

Summary Table. Checkpoints are mechanisms that impose delays in the cell cycle in response to DNA damage or defects in DNA replication, to ensure that mitotic transmission is error-free. Failure to delay the cell cycle in the presence of damage converts an easily reparable DNA lesion into one far more deleterious, provoking genomic instability or cell death. This figure shows a summary of our current knowledge about the DNA damage checkpoints in yeast. Genetic analysis of the pathway has allowed classification of its components into