RIPAb+ SMN - RIP Validated Antibody and Primer Set

The survival of motor neurons (SMN) protein is essential for the biogenesis of small nuclear RNA (snRNA)-ribonucleoproteins (snRNPs), the major components of the pre-mRNA splicing machinery. Though it is ubiquitously expressed, SMN deficiency causes the motor neuron degenerative disease spinal muscular atrophy (SMA). SMN deficiency has unexpected cell type-specific effects on the repertoire of snRNAs and mRNAs. It alters the stoichiometry of snRNAs and causes widespread pre-mRNA splicing defects in numerous transcripts of diverse genes, preferentially those containing a large number of introns, in SMN-deficient mouse tissues. The SMN complex plays a role in RNA metabolism and in splicing regulation.

~35 kDa was observed; however, the calculated molecular weight is 31.849 kDa

Epitope: Unknown

Histidine-tagged recombinant protein corresponding to human SMN.

RNA Binding Protein Immunoprecipitation:
Representative lot data.
RIP Lysate prepared from HeLa cells (~2 X 10E7 cell equivalents per IP) were subjected to immunoprecipitation using either 5 µg of a normal mouse IgG or 5 µg of Anti-SMN antibody and the Magna RIP^® RNA-Binding Protein Immunoprecipitation Kit (Cat. # 17-700).
Successful immunoprecipitation of SMN-associated RNA was verified by qPCR using primers specific for human collagen alpha type 1 mRNA (Please see figures).
Please refer to the Magna RIP (Cat. # 17-700) or EZ-Magna RIP (Cat. # 17-701) protocol for experimental details.

Western Blot Analysis:
Representative lot data.
HeLa cell lysate was resolved by electrophoresis, transferred to PVDF membranes and probed with anti-SMN at a 0.5 µg/mL dilution.
Proteins were visualized using a goat anti-mouse IgG conjugated to HRP using a chemiluminescence detection system.
Arrow indicates SMN (~35 kDa) (Please see figures).

Immunocytochemistry Analysis:
Representative lot data.
A 1:500 dilution from a representative lot detected SMN in HeLa cells (Please see figures).

Research Category
Epigenetics & Nuclear Function

Research Sub Category
RNA Metabolism & Binding Proteins

This RIPAb+ SMN -RIP Validated Antibody & Primer Set conveniently includes the antibody & the specific control PCR primers.

10 assays per set. Recommended use: ~5 μg of antibody per RIP (dependent upon biological context).

Anti-SMN (Mouse Monoclonal). One vial containing 50 µg of protein G purified monoclonal IgG1ĸ in buffer containing 0.1 M Tris-glycine, 150 mM NaCl, pH 7.4, 0.05% sodium azide before the addition of 30% glycerol. Store at -20°C.

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

RIP Primers, U1snRNA. One vial containing 75 μL of 5 μM of each primer specific for the cDNA of U1 snRNP. Store at -20°C.
FOR: GGG AGA TAC CAT GAT CAC GAA GGT
REV: CCA CAA ATT ATG CAG TCG AGT TTC CC

Format: Purified

Protein G Purified

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.

RNA Binding Protein Immunoprecipitation:
RIP Lysate prepared from HeLa cells (~2 X 10E7 cell equivalents per IP) were subjected to immunoprecipitation using either 5 µg of a normal mouse IgG or 5 µg of Anti-SMN antibody and the Magna RIP^® RNA-Binding Protein Immunoprecipitation Kit (Cat. # 17-700).
Successful immunoprecipitation of SMN-associated RNA was verified by qPCR using RIP Primers U1snRNA (Please see figures).
Please refer to the Magna RIP (Cat. # 17-700) or EZ-Magna RIP (Cat. # 17-701) protocol for experimental details.

Control
Includes negative control mouse IgG antibody and control primers specific for the cDNA of human U1snRNP.

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