RIPAb+ Lin28 Kit

• 产品编号：merck-m-03-105      品牌：millipore       原厂货号：03-105
• 产品分类：抗体 > 一抗 > 蛋白特异性一抗
• 应用分类：

描述：

Description:RIPAb+ Lin28 - RIP Validated Antibody and Primer Set.

Overview:RIPAb+ antibodies are evaluated using the RNA Binding Protein Immunoprecipitation (RIP) assay. Each RIPAb+ antibody set includes quantitative RT-PCR control primers (RIP Primers) to biologically validate your IP results by successfully coprecipitating a specific RNA target (where such a specific target is known). The qPCR protocol and primer sequences are provided, allowing researchers to validate RIP protocols when using the antibody in their experimental context. Each set also includes a negative control antibody to ensure specificity of the RIP reaction.
The RIPAb+ Lin28 set includes the Lin28 (Zinc Finger CCHC domain-containing protein 1) antibody, a negative control antibody (purified rabbit IgG), and positive qPCR primers which amplify a 68 bp fragment of the human cDNA of CDK4. The Lin28 and negative control antibodies are supplied in a scalable "per RIP" reaction size and can be used to functionally validate the precipitation of Lin28-associated RNA.

Background Information:Lin28 is a developmentally regulated RNA binding protein (RBP). It is down regulation is mediated in part by Lin4 and miR-125b, the mammalian micro-RNA homolog of Lin4. It is known to be negatively regulated by retinoic acid and act as a translational enhancer by driving specific mRNAs to polysomes and thus increasing the efficiency of protein synthesis by increasing the number of initiation events per mRNA molecule. Lin28 is necessary and sufficient for selectively blocking the cleavage processing of pri-let-7 miRNAs in embryonic cells (Viswanathan, 2008). Lin28 is a negative regulator of miRNA biogenesis and may play a role in blocking miRNA-mediated differentiation in stem cells and in certain cancers (Viswanathan, 2008). If this degradation does not occur, then normal development of various somatic cells fails. The mechanism by which miR-125b represses Lin28 in mammalian cells appears to involve reductions not only in translational efficiency, but also in mRNA abundance (Wu, 2005). Lin28 has shown to be essential for skeletal muscle differentiation through the translational upregulation of IGF2 expression. It is known to be expressed in testis, placenta, and embryonic stem (ES) cells.

Presentation:Anti-Lin28 (Rabbit Polyclonal IgG). One vial containing 50 µL of rabbit serum. Store at -20°C.

Normal Rabbit IgG. One vial containing 125 μg of purified rabbit IgG in 125 μL storage buffer containing 0.1% sodium azide. Store at -20°C.

RIP Primers, CDK4. One vial containing 75 μL of 5 μM of each primer specific for the cDNA of human CDK4. Store at -20°C.
FOR: ATG TTG TCC GGC TGA TGG A
REV: CAC CAG GGT TAC CTT GAT CTC C

Key Applications

• Western Blotting
• RNA Binding Protein Immunoprecipitation (RIP)
• Immunoprecipitation

原厂资料：

Description:RIPAb+ Lin28 - RIP Validated Antibody and Primer Set.

Overview:RIPAb+ antibodies are evaluated using the RNA Binding Protein Immunoprecipitation (RIP) assay. Each RIPAb+ antibody set includes quantitative RT-PCR control primers (RIP Primers) to biologically validate your IP results by successfully coprecipitating a specific RNA target (where such a specific target is known). The qPCR protocol and primer sequences are provided, allowing researchers to validate RIP protocols when using the antibody in their experimental context. Each set also includes a negative control antibody to ensure specificity of the RIP reaction.
The RIPAb+ Lin28 set includes the Lin28 (Zinc Finger CCHC domain-containing protein 1) antibody, a negative control antibody (purified rabbit IgG), and positive qPCR primers which amplify a 68 bp fragment of the human cDNA of CDK4. The Lin28 and negative control antibodies are supplied in a scalable "per RIP" reaction size and can be used to functionally validate the precipitation of Lin28-associated RNA.

Background Information:Lin28 is a developmentally regulated RNA binding protein (RBP). It is down regulation is mediated in part by Lin4 and miR-125b, the mammalian micro-RNA homolog of Lin4. It is known to be negatively regulated by retinoic acid and act as a translational enhancer by driving specific mRNAs to polysomes and thus increasing the efficiency of protein synthesis by increasing the number of initiation events per mRNA molecule. Lin28 is necessary and sufficient for selectively blocking the cleavage processing of pri-let-7 miRNAs in embryonic cells (Viswanathan, 2008). Lin28 is a negative regulator of miRNA biogenesis and may play a role in blocking miRNA-mediated differentiation in stem cells and in certain cancers (Viswanathan, 2008). If this degradation does not occur, then normal development of various somatic cells fails. The mechanism by which miR-125b represses Lin28 in mammalian cells appears to involve reductions not only in translational efficiency, but also in mRNA abundance (Wu, 2005). Lin28 has shown to be essential for skeletal muscle differentiation through the translational upregulation of IGF2 expression. It is known to be expressed in testis, placenta, and embryonic stem (ES) cells.

Presentation:Anti-Lin28 (Rabbit Polyclonal IgG). One vial containing 50 µL of rabbit serum. Store at -20°C.

Normal Rabbit IgG. One vial containing 125 μg of purified rabbit IgG in 125 μL storage buffer containing 0.1% sodium azide. Store at -20°C.

RIP Primers, CDK4. One vial containing 75 μL of 5 μM of each primer specific for the cDNA of human CDK4. Store at -20°C.
FOR: ATG TTG TCC GGC TGA TGG A
REV: CAC CAG GGT TAC CTT GAT CTC C

Key Applications

• Western Blotting
• RNA Binding Protein Immunoprecipitation (RIP)
• Immunoprecipitation

注意事项：

Application Notes

RNA Binding Protein Immunoprecipitation:
RIP Lysate prepared from NTera-2 embryonic carcinoma cells (2 X 10⁷ cell equivalents per IP) were subjected to immunoprecipitation using 5 µl of either a normal rabbit IgG, or Anti-Lin28 antibody and the Magna RIP™ Kit (Cat. # 17-700). 
Successful immunoprecipitation of Lin28-associated RNA was verified by qPCR using primers for U1snRNA as a negative control and positive control RIP Primers CDK4 (Please see figures). Data is presented as percent input of each IP sample relative to input total RNA for each amplicon and RIP sample as indicated. Please refer to the Magna RIP™ (Cat. # 17-700) or EZ-Magna RIP™ (Cat. # 17-701) protocol for experimental details.

Immunoprecipitation from RIP lysate:
RIP lysate from NTera-2 embryonic carcinoma cells (2 X 10⁶ cell equivalents per IP) was subjected to immunoprecipitation using 0.5 µL of either a normal rabbit IgG or Anti-Lin28 antibody. Precipitated proteins were resolved by electrophoresis, transferred to nitrocellulose and probed with anti-Lin28 (1.0 µg/mL). Proteins were visualized using One-Step™ IP-Western kit (GenScript Cat. # L00231) (Please see figures).

Storage Conditions

Stable for 1 year at -20°C from date of receipt. Store at -80°C for longer term storage. Handling Recommendations: Upon first thaw, 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.