Cell Signaling Technology

Product Pathways - siRNA

SignalSilence® p38α MAPK siRNA I (Mouse Specific) #6417

Applications Reactivity
Transfection M

Reactivity Key:  M=Mouse
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Western Blotting

Western Blotting

Western blot analysis of extracts from NIH/3T3 cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-), SignalSilence® p38α MAPK siRNA I (Mouse Specific) (+) or SignalSilence® p38α MAPK siRNA II (Mouse Specific) #6418 (+), using p38α MAPK Antibody #9218 (upper) or α-Tubulin (11H10) Rabbit mAb #2125 (lower). The p38α MAPK Antibody confirms silencing of p38α MAPK expression, while the α-Tubulin (11H10) Rabbit mAb is used as a loading control.

Description

SignalSilence® p38α MAPK siRNA I (Mouse Specific) from Cell Signaling Technology (CST) allows the researcher to specifically inhibit p38α MAPK expression using RNA interference, a method whereby gene expression can be selectively silenced through the delivery of double stranded RNA molecules into the cell. All SignalSilence® siRNA products from CST are rigorously tested in-house and have been shown to reduce target protein expression by western analysis.

Quality Control

Oligonucleotide synthesis is monitored base by base through trityl analysis to ensure appropriate coupling efficiency. The oligo is subsequently purified by affinity-solid phase extraction. The annealed RNA duplex is further analyzed by mass spectrometry to verify the exact composition of the duplex. Each lot is compared to the previous lot by mass spectrometry to ensure maximum lot-to-lot consistency.

Directions for Use

CST recommends transfection with 100 nM p38α MAPK siRNA I (Mouse Specific) 48 to 72 hours prior to cell lysis. For transfection procedure, follow protocol provided by the transfection reagent manufacturer. Please feel free to contact CST with any questions on use.

Each vial contains the equivalent of 100 transfections, which corresponds to a final siRNA concentration of 100 nM per transfection in a 24-well plate with a total volume of 300 μl per well.

Background

p38 MAP kinase (MAPK), also called RK (1) or CSBP (2), is the mammalian orthologue of the yeast HOG kinase that participates in a signaling cascade controlling cellular responses to cytokines and stress (1-4). Four isoforms of p38 MAPK, p38α, β, γ (also known as Erk6 or SAPK3), and δ (also known as SAPK4) have been identified. Similar to the SAPK/JNK pathway, p38 MAPK is activated by a variety of cellular stresses including osmotic shock, inflammatory cytokines, lipopolysaccharide (LPS), UV light, and growth factors (1-5). MKK3, MKK6, and SEK activate p38 MAPK by phosphorylation at Thr180 and Tyr182. Activated p38 MAPK has been shown to phosphorylate and activate MAPKAP kinase 2 (3) and to phosphorylate the transcription factors ATF-2 (5), Max (6), and MEF2 (5-8).SB203580 (4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-imidazole) is a selective inhibitor of p38 MAPK. This compound inhibits the activation of MAPKAPK-2 by p38 MAPK and subsequent phosphorylation of HSP27 (9). SB203580 inhibits p38 MAPK catalytic activity by binding to the ATP-binding pocket, but does not inhibit phosphorylation of p38 MAPK by upstream kinases (10).

  1. Rouse, J. et al. (1994) Cell 78, 1027-1037.
  2. Han, J. et al. (1994) Science 265, 808-811.
  3. Lee, J.C. et al. (1994) Nature 372, 739-746.
  4. Freshney, N.W. et al. (1994) Cell 78, 1039-1049.
  5. Raingeaud, J. et al. (1995) J. Biol. Chem. 270, 7420-7426.
  6. Zervos, A.S. et al. (1995) Proc. Natl. Acad. Sci. USA 92, 10531-10534.
  7. Zhao, M. et al. (1999) Mol. Cell. Biol. 19, 21-30.
  8. Yang, S.H. et al. (1999) Mol. Cell. Biol. 19, 4028-4038.
  9. Cuenda, A. et al. (1995) FEBS Lett 364, 229-33.
  10. Kumar, S. et al. (1999) Biochem Biophys Res Commun 263, 825-31.

Application References

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Companion Products

Limited Use Label License, RNA interference: This product is licensed under European Patent 1144623 and foreign equivalents from Ribopharma AG, Kulmbach, Germany and is provided only for use in non-commercial research specifically excluding use (a) in drug discovery or drug development, including target identification or target validation, by or on behalf of a commercial entity, (b) for contract research or commercial screening services, (c) for the production or manufacture of siRNA-related products for sale, or (d) for the generation of commercial databases for sale to Third Parties. Information about licenses for these and other commercial uses is available from Ribopharma AG, Fritz-Hornschuch-Str. 9, D-95326 Kulmbach, Germany.

For Research Use Only. Not For Use In Diagnostic Procedures.

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