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03-191
Sigma-AldrichRIPAb+ Upf1 - RIP Validated Antibody and Primer Set
This RIPAb+ Upf1 -RIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers.
More>>This RIPAb+ Upf1 -RIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers. Less<<
SDB (Sicherheitsdatenblätter), Analysenzertifikate und Qualitätszertifikate, Dossiers, Broschüren und andere verfügbare Dokumente.
RIPAb+ Upf1 - RIP Validated Antibody and Primer Set
Overview
RIPAb+ antibodies are evaluated using the RNA Binding Protein Immunoprecipitation (RIP) assay. Each RIPAb+ antibody set includes a negative control antibody to ensure specificity of the RIP reaction and is verified for the co-immunoprecipitation of RNA associated specifically with the immunoprecipitated RNA binding protein of interest. Where appropriate, the RIPAb+ set also includes quantitative RT-PCR control primers (RIP Primers) to biologically validate your IP results by successfully co-precipitating the specific RNA targets, such as messenger RNAs. The qPCR protocol and primer sequences are provided, allowing researchers to validate RIP protocols when using the antibody in their experimental context. If a target specific assay is not provided, the RIPAb+ kit is validated using an automated microfluidics-based assay by enrichment of detectable RNA over control immunoprecipitation. Upf1 is part of a post-splicing multiprotein complex involved in both mRNA nuclear export and mRNA surveillance. Upf1 is located only in the cytoplasm. When translation ends, it interacts with the protein that is a functional homolog of yeast Upf2p to trigger mRNA decapping.
Alternate Names
ATP-dependent helicase RENT1
Nonsense mRNA reducing factor 1
UP Frameshift 1
UPF1 regulator of nonsense transcripts homolog (yeast)
Up-frameshift suppressor 1 homolog
delta helicase
regulator of nonsense transcripts 1
up-frameshift mutation 1 homolog
yeast Upf1p homolog
Background Information
Upf1 is part of a post-splicing multiprotein complex involved in both mRNA nuclear export and mRNA surveillance. mRNA surveillance detects exported mRNAs with truncated open reading frames and initiates nonsense-mediated mRNA decay (NMD). When translation ends upstream from the last exon-exon junction, this triggers NMD to degrade mRNAs containing premature stop codons. Upf1 is located only in the cytoplasm. When translation ends, it interacts with the protein that is a functional homolog of yeast Upf2p to trigger mRNA decapping.
References
Product Information
Format
Purified
Control
Includes negative control normal rabbit IgG antibody and control primers specific for the cDNA of human ARF1.
Presentation
Anti-Upf1 (Rabbit Polyclonal). One vial containing 70 μL of purified rabbit polyclonal antibody in buffer containing 0.1 M Tris-Glycine (pH 7.4), 150 mM NaCl with 0.05% sodium azide and 30% glycerol. Store at -20°C.
Normal Rabbit IgG. One vial containing 125 µg of rabbit IgG in 125 µL of storage buffer containing 0.05% sodium azide. Store at -20°C.
RIP Primers, ARF1. One vial containing 75 μL of 5 μM of each primer specific for human ARF1. Store at -20°C. FOR: GAC CAC GAT CCT CTA CAA GC REV: TCC CAC ACA GTG AAG CTG ATG
This RIPAb+ Upf1 -RIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers.
Key Applications
RNA Binding Protein Immunoprecipitation (RIP)
Western Blotting
Immunoprecipitation
Application Notes
Immunoprecipitation from RIP lysate: Representative lot data. RIP lysate from HeLa cells (~2 X 10E7 cell equivalents per IP) was subjected to immunoprecipitation using either 7 µL of a normal rabbit IgG or 7 µL of Anti-Upf1 antibody. Ten percent of the precipitated proteins (lane 1: rabbit IgG, lane 2: Upf1) were resolved by electrophoresis, transferred to PVDF and probed with anti-Upf1 antibody (1:2000). Proteins were visualized using One-Step™ IP-Western kit (GenScript Cat. # L00232) Arrow indicates Upf1 (~125 kDa) (Please see figures).
Western Blot Analysis: Representative lot data. U2OS cell lysate was resolved by electrophoresis, transferred to PVDF membrane and probed with anti-Upf1 (0.5 μg/mL). Proteins were visualized using a donkey anti-rabbit secondary antibody conjugated to HRP and a chemiluminescence detection system. Arrow indicates Upf1 (~125 kDa) (Please see figures).
Biological Information
Immunogen
KLH-conjugated synthetic linear peptide corresponding to Upf1.
Host
Rabbit
Specificity
This antibody detects Upf1.
Species Reactivity
Bovine
Human
Mouse
Rat
Canine
Species Reactivity Note
Demonstrated to react with human. Predicted to react with mouse, rat, bovine and canine based on sequence homology.
This gene encodes a protein that is part of a post-splicing multiprotein complex involved in both mRNA nuclear export and mRNA surveillance. mRNA surveillance detects exported mRNAs with truncated open reading frames and initiates nonsense-mediated mRNA decay (NMD). When translation ends upstream from the last exon-exon junction, this triggers NMD to degrade mRNAs containing premature stop codons. This protein is located only in the cytoplasm. When translation ends, it interacts with the protein that is a functional homolog of yeast Upf2p to trigger mRNA decapping. Use of multiple polyadenylation sites has been noted for this gene. [provided by RefSeq].
FUNCTION: Part of a post-splicing multiprotein complex. Involved in nonsense-mediated decay (NMD) as part of the SMG1C complex, a mRNA surveillance complex that recognizes and degrades mRNAs containing premature translation termination codons (PTCs). The complex probably acts by associating with ribosomes during tranlation termination on mRNPs. If an exon junction complex (EJC) is located 50-55 or more nucleotides downstream from the termination codon, RENT1 is phosphorylated by SMG1, triggering nonsense-mediated decay (NMD). Essential for embryonic viability.
SUBUNIT STRUCTURE: Found in a complex with RENT1, RENT2, RENT3A and RENT3B. Found in a complex with PARN. Found in a post-splicing complex with SMG1, NXF1, RBM8A, RENT1, RENT2, RENT3A, RENT3B and RNPS1. Found in a mRNA decay complex with EXOSC2, EXOSC4, EXOSC10, PARN, XRN1, DCP2, RENT1, RENT2 and RENT3B. Interacts with EST1A and RENT2. Component of the SMG1C complex, at least composed of SMG1, SMG8 and SMG9. The SMG1C complex is then recruited on premature translation termination codons (PTCs) to form the ribosome:SURF complex, at least composed of ERF1, ERF3 (ERF3A or ERF3B), EEF2, UPF1/RENT1, SMG1, SMG8 and SMG9. Interacts (when hyperphosphorylated) with PNRC2.
SUBCELLULAR LOCATION: Cytoplasm. Cytoplasm › P-body. Note: Hyperphosphorylated form is targeted to the P-body, while unphosphorylated protein is distributed throughout the cytoplasm.
TISSUE SPECIFICITY: Ubiquitous.
DOMAIN: The [ST]-Q motif constitutes a recognition sequence for kinases from the PI3/PI4-kinase family.
PTM: Phosphorylated by SMG1; required for formation of mRNA surveillance complexes. Phosphorylated upon DNA damage, probably by ATM or ATR.
SEQUENCE SIMILARITIES: Belongs to the DNA2/NAM7 helicase family.
Contains 1 C2H2-type zinc finger.
Molecular Weight
~125 kDa
Physicochemical Information
Dimensions
Materials Information
Toxicological Information
Safety Information according to GHS
Safety Information
Product Usage Statements
Quality Assurance
RNA Binding Protein Immunoprecipitation: RIP Lysate prepared from HeLa cells (2 X 10E7 cell equivalents per IP) were subjected to immunoprecipitation using either 7 µL of a normal rabbit IgG or 7 µL Anti-Upf1 antibody and the Magna RIP™ RNA-Binding Protein Immunoprecipitation Kit (Cat. # 17-700). Successful immunoprecipitation of Upf1-associated RNA was verified by qPCR using RIP Primers ARF1 (Please see figures). Please refer to the Magna RIP™ (Cat. # 17-700) or EZ-Magna RIP™ (Cat. # 17-701) protocol for experimental details.
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
Storage Conditions
Stable for 1 year at -20°C from date of receipt. Handling Recommendations: Upon receipt, and prior to removing the cap, centrifuge the vial and gently mix the solution. Aliquot into microcentrifuge tubes and store at -20°C. Avoid repeated freeze/thaw cycles, which may damage IgG and affect product performance. Note: Variability in freezer temperatures below -20°C may cause glycerol containing solutions to become frozen during storage.
Packaging Information
Material Size
10 assays
Material Package
10 assays per set. Recommended use: ~7 μL of antibody per RIP (dependent upon biological context).
Transport Information
Supplemental Information
Specifications
Global Trade Item Number
Bestellnummer
GTIN
03-191
04053252518928
Documentation
RIPAb+ Upf1 - RIP Validated Antibody and Primer Set SDB
Eukaryotic RNAs with premature termination codons (PTCs) are eliminated by nonsense-mediated decay (NMD). While human nonsense RNA degradation can be initiated either by an endonucleolytic cleavage event near the PTC or through decapping, the individual contribution of these activities on endogenous substrates has remained unresolved. Here we used concurrent transcriptome-wide identification of NMD substrates and their 5'-3' decay intermediates to establish that SMG6-catalyzed endonucleolysis widely initiates the degradation of human nonsense RNAs, whereas decapping is used to a lesser extent. We also show that a large proportion of genes hosting snoRNAs in their introns produce considerable amounts of NMD-sensitive splice variants, indicating that these RNAs are merely by-products of a primary snoRNA production process. Additionally, transcripts from genes encoding multiple snoRNAs often yield alternative transcript isoforms that allow for differential expression of individual coencoded snoRNAs. Based on our findings, we hypothesize that snoRNA host genes need to be highly transcribed to accommodate high levels of snoRNA production and that the expression of individual snoRNAs and their cognate spliced RNA can be uncoupled via alternative splicing and NMD.
