Cell Signaling Technology

Product Pathways - MAPK Signaling

SignalSilence® Pool p38 MAPK siRNA Kit #6385

Kit Includes

SignalSilence® Pool p38 MAP Kinase siRNA #6386
pre-validated heterogeneous duplex siRNA guaranteed to specifically inhibit p38 MAP Kinase expression in specified cell lines.
SignalSilence® Control siRNA (Fluorescein Conjugate) #6201
a non-targeted negative control duplex designed to monitor transfection efficiency using microscopy and to monitor siRNA specificity.Kit contains enough for 4(S) or 10 (L) transfections.
Targeted p38 MAP Kinase Antibody #9212
to confirm silencing of p38 MAP Kinase expression.
Non-targeted ATP-Citrate Lyase Antibody #4332
to control for protein loading and p38 MAP Kinase siRNA specificity.
*Transfection Reagent
designed specifically for highly efficient siRNA delivery in mammalian cells. Transfection reagent is supplied by Mirus.

*Store at 4° (packaged separately)

Western Blotting

Western Blotting

Western blot analysis of extracts from HeLa cells transfected with 20 nM control siRNA #6201, or 5, 20 nM pool p38 MAP Kinase siRNA, using p38 MAP Kinase Antibody #9212 (upper), p44/42 MAP Kinase Antibody #9102 (middle) or SAPK/JNK (56G8) Rabbit mAb #9258 (lower) in combination with ATP-Citrate Lyase Antibody #4332. The p38 MAP Kinase Antibody confirms silencing of p38 MAP Kinase expression, and ATP-Citrate Lyase Antibody is used to control for loading and specificity of pool p38 MAP Kinase siRNA. The experiment also demonstrates specifically that the pool p38 MAPK siRNA does not interfere with the expression of p44/42 MAPK or SAPK/JNK.

Fluorescent Detection

Fluorescent Detection

Fluorescent detection of SignalSilence® Control siRNA (Fluorescein Conjugate) #6201 in living HeLa cells 24 hours post-transfection, demonstrating nearly 100% transfection efficiency.

Description

SignalSilence® Pool p38 MAP Kinase siRNA Kit from Cell Signaling Technology allows the researcher to specifically inhibit p38 MAP Kinase expression. The kit utilizes RNA interference, a method in which gene expression can be selectively silenced through the delivery of double stranded RNA molecules into the cell. The p38 MAP Kinase siRNA is a hetergeneous mixture of 21-22 bp siRNAs. A 418 bp cDNA template of mouse p38 MAP Kinase was transcribed by T7 RNA polymerase to create double-stranded RNA (dsRNA). RNase III was used to cleave the dsRNA in the presence of Mn2+ buffer to 21-22 bp siRNA. All SignalSilence® siRNA kits are rigorously tested in-house and have been shown to reduce protein expression in specified cell lines. SignalSilence® Pool p38 MAP Kinase Kit includes Pool p38 MAP Kinase siRNA, a target-specific p38 MAP Kinase Antibody to confirm the silencing of p38 MAP Kinase expression and a non-target antibody (ATP-Citrate Lyase) to control for loading and monitor specificity of p38 MAP Kinase siRNA. Additionally, a fluorescein-labeled non-targeted siRNA control allows the user to monitor transfection efficiency and siRNA specificity. The kit also contains a transfection reagent designed solely for efficient siRNA delivery in mammalian cells.

Directions for Use

CST recommends transfection with 20 nM pool p38 MAP Kinase siRNA 48 to 72 hours prior to cell lysis. See Protocol for transfection procedure.

Background

p38 MAP kinase (MAPK), also called RK (1) or CSBP (2), is the mammalian orthologue of the yeast HOG kinase which participates in a signaling cascade controlling cellular responses to cytokines and stress (1-4). Four isoforms of p38 MAP kinase, 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 MAP kinase is activated by a variety of cellular stresses including osmotic shock, inflammatory cytokines, lipopolysaccharides (LPS), UV light and growth factors (1-5). MKK3, MKK6 and SEK activate p38 MAP kinase by phosphorylation at Thr180 and Tyr182. Activated p38 MAP kinase 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).

  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.

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.

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