Cell Signaling Technology

Product Pathways - Cell Cycle / Checkpoint

SignalSilence® CDK8 siRNA I #6438

Applications Reactivity
Transfection H (M) (R)

Reactivity Key:  H=Human  M=Mouse  R=Rat
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 293 cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-) or SignalSilence® CDK8 siRNA I (+), using CDK8 (G398) Antibody #4101 and α-Tubulin (11H10) Rabbit mAb #2125. The CDK8 (G398) Antibody confirms silencing of CDK8 expression, while the α-Tubulin (11H10) Rabbit mAb is used as a loading control.

Description

SignalSilence® CDK8 siRNA I from Cell Signaling Technology (CST) allows the researcher to specifically inhibit CDK8 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 SignalSilence® CDK8 siRNA I 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

The mammalian Mediator Complex is a multi-subunit protein complex that couples specific transcriptional regulators to RNA polymerase II (Pol II) and the basal transcription machinery. Interactions between distinct Mediator subunits and transcription factors allow for specific gene regulation (reviewed in 1).Mediator complex interactions control various biological processes including insulin signaling (2), NF-κB-dependent signaling (3), stem cell pluripotency and self renewal (4,5), and proliferation of colon cancer cells (6,7).CDK8/Cyclin C, along with Med12 and Med13, constitute a subcomplex within the Mediator Complex thought to act as a molecular switch, inhibiting Pol II recruitment and transcription initiation (8,9). Expression of CDK8 abrogates E2F-1-dependent inhibition of β-catenin activity in colon cancer cells (9). High levels of CDK8 coincide with high β-catenin-dependent transcription in colon cancer cells, and their proliferation can be inhibited by suppressing CDK8 expression (8).

  1. Malik, S. and Roeder, R.G. (2005) Trends Biochem Sci 30, 256-63.
  2. Wang, W. et al. (2009) Dev Cell 16, 764-71.
  3. van Essen, D. et al. (2009) PLoS Biol 7, e73.
  4. Tutter, A.V. et al. (2009) J Biol Chem 284, 3709-18.
  5. Varelas, X. et al. (2008) Nat Cell Biol 10, 837-48.
  6. Firestein, R. et al. (2008) Nature 455, 547-51.
  7. Morris, E.J. et al. (2008) Nature 455, 552-6.
  8. Knuesel, M.T. et al. (2009) Mol Cell Biol 29, 650-61.
  9. Knuesel, M.T. et al. (2009) Genes Dev 23, 439-51.

Application References

Have you published research involving the use of our products? If so we'd love to hear about it. Please let us know!

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.

Products