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S7813 CpG WIZ® RASSF1A Amplification Kit

The components of the CpG WIZ RASSF1A Amplification Kit include those required for PCR amplification after bisulfite modification of DNA samples. Sufficient reagents are provided to analyze 25 samples.

S7813
25 assays  
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      Overview

      Replacement Information
      Description
      Catalogue NumberS7813
      Brand Family Chemicon®
      Trade Name
      • CpG Wiz
      • Chemicon
      DescriptionCpG WIZ® RASSF1A Amplification Kit
      OverviewMethylation of cytosines located 5' to guanosine is known to have a profound effect on the expression of several eukaryotic genes (Bird, 1992). In normal cells, methylation occurs predominantly in CG-poor regions, while CG-rich areas, called CpG islands, remain unmethylated. Aberrant methylation of normally unmethylated CpG islands has been documented as a relatively frequent event in immortalized and transformed cells (Antequera, 1990) and has been associated with transcriptional inactivation of defined tumor suppressor genes in human cancers (Herman, 1994; Merlo, 1995). The Ras association domain family 1 (RASSF1A) gene exhibits characteristic hypermethylation.



      Methylation-specific PCR (MSP) is a new technology for sensitive detection of abnormal gene methylation utilizing small amounts of DNA (Herman, 1996). This process employs an initial bisulfite reaction to modify the DNA, followed by PCR amplification with specific primers designed to distinguish methylated from unmethylated DNA.
      Materials Required but Not DeliveredEquipment and Supplies

      a. Microcentrifuge tubes for PCR amplification

      b. Aerosol-resistant pipette tips

      c. Thermocycler

      d. Gel electrophoresis apparatus (vertical or horizontal)

      e. Power Supply

      f. 302 nm UV transilluminator, camera and film

      Reagents

      a. 2.5 mM dNTP mix (2.5 mM of each nucleotide)

      b. "Hot start" Taq polymerase

      c. "Hot start" PCR reagents (see Sec. II. Protocols).

      d. Reagents for gel electrophoresis (1X TBE and 2% agarose, 10% acrylamide, or suitable high resolution agarose)

      e. DNA markers (size range 100-300 bp)

      f. Ethidium bromide (10 mg/mL)

      g. Gel-loading solution / Loading Dye

      h. Bisulfite Modified DNA (CpGenome™ DNA Modification Kit, S7820)
      References
      Product Information
      Components
      • The components of the CpG WIZ® RASSF1A Amplification Kit include those required for PCR amplification after bisulfite modification of DNA samples. Sufficient reagents are provided to analyze 25 samples with appropriate controls.
      • U Primer Set5 μM each primer (25X) 35 μL (neutral cap) 90566 -15°C to -25°C
      • M Primer Set5 μM each primer (25X) 35 μL (red cap) 90567 -15°C to -25°C
      • W Primer Set5 μM each primer (25X) 35 μL (green cap) 90568 -15°C to -25°C
      • U control DNA0.1 μg/μL 50 μL (white cap) 90393 -15°C to -25°C
      • M control DNA0.1 μg/μL 50 μL (red cap) 90394 -15°C to -25°C
      • W control DNA0.05 μg/μL 50 μL (green cap) 90395 -15°C to -25°C
      • Universal 10X PCR Buffer 265 μL (blue cap) 90396 -15°C to -25°C
      Quality LevelMQ100
      Applications
      ApplicationThe components of the CpG WIZ RASSF1A Amplification Kit include those required for PCR amplification after bisulfite modification of DNA samples. Sufficient reagents are provided to analyze 25 samples.
      Key Applications
      • PCR
      Application NotesPrinciples of the Technique

      Use of either the CpGenome™ DNA Modification Kit (Cat. No. S7820) or the CpGENOME™ Fast DNA Modification Kit (Cat. No. S7824) facilitates the initial bisulfite reactions, while the CpG WIZ® RASSF1A Amplification Kit contains the reagents required for the gene-specific PCR amplification reactions.
      Biological Information
      Species Reactivity
      • Human
      Entrez Gene Number
      Entrez Gene SummaryThis gene encodes a protein similar to the RAS effector proteins. Loss or altered expression of this gene has been associated with the pathogenesis of a variety of cancers, which suggests the tumor suppressor function of this gene. The inactivation of this gene was found to be correlated with the hypermethylation of its CpG-island promoter region. The encoded protein was found to interact with DNA repair protein XPA. The protein was also shown to inhibit the accumulation of cyclin D1, and thus induce cell cycle arrest. Seven alternatively spliced transcript variants of this gene encoding distinct isoforms have been reported.
      Gene Symbol
      • RASSF1
      • REH3P21
      • RDA32
      • WUGSC:H_LUCA12.5
      • RASSF1A
      • 123F2
      • NORE2A
      UniProt Number
      UniProt SummaryFUNCTION: SwissProt: Q9NS23 # Potential tumor suppressor. Required for death receptor- dependent apoptosis. Mediates activation of STK4 during Fas- induced apoptosis. When associated with MOAP1, promotes BAX conformational change and translocation to mitochondrial membranes in response to TNF and TNFSF10 stimulation. Isoform A interacts with CDC20, an activator of the anaphase-promoting complex, APC, resulting in the inhibition of APC activity and mitotic progression. Inhibits proliferation by negatively regulating cell cycle progression at the level of G1/S-phase transition by regulating accumulation of cyclin D1 protein. Isoform C has been shown not to perform these roles, no function has been identified for this isoform.
      SIZE: 344 amino acids; 39219 Da
      SUBUNIT: Interacts with MAP1S and XPA. Binds to the N-terminal of CDC20 during prometaphase. Binds to STK3 and STK4. Isoform A interacts with MOAP1. Recruited to the TNFRSF1A and TNFRSF10A complexes in response to their respective cognate ligand, after internalization. Can self-associate. Isoform A interacts with RSSF5 and probably associates with HRAS1 via a RSSF1 isoform A- RSSF5 heterodimer.
      SUBCELLULAR LOCATION: Isoform A: Cytoplasm, cytoskeleton. Centrosome. Spindle. Note=Localizes to cytoplasmic microtubules during interphase, to bipolar centrosomes associated with microtubules during prophase, to spindle fibers and spindle poles at metaphase and anaphase, to the midzone during early telophase, and to the midbody in late telophase in cells. & Isoform C: Nucleus. Note=Predominantly nuclear.TISSUE SPECIFICITY: Isoform A and isoform C are ubiquitously expressed in all tissues tested, however isoform A is absent in many corresponding cancer cell lines. Isoform B is mainly expressed in hematopoetic cells.
      SIMILARITY: SwissProt: Q9NS23 ## Contains 1 phorbol-ester/DAG-type zinc finger. & Contains 1 Ras-associating domain. & Contains 1 SARAH domain.
      Physicochemical Information
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Usage Statement
      • Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
      Storage and Shipping Information
      Packaging Information
      Material Size25 assays
      Transport Information
      Supplemental Information
      Specifications
      Global Trade Item Number
      Catalog Number GTIN
      S7813 04053252663529

      Documentation