Cell Signaling Technology

Product Pathways - Cell Cycle / Checkpoint

SignalSilence® Rb siRNA II #6542

Applications Reactivity
Transfection H

Reactivity Key:  H=Human
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 HeLa cells, transfected with 100 nM SignalSilence® Control siRNA (Fluorescein Conjugate) #6201 (-), SignalSilence® Rb siRNA I #6451 (+) or SignalSilence® Rb siRNA II (+), using Rb (4H1) Mouse mAb #9309 and α-Tubulin (11H10) Rabbit mAb #2125. The Rb (4H1) Mouse mAb confirms silencing of Rb expression, while the α-Tubulin (11H10) Rabbit mAb is used to control for loading and specificity of Rb siRNA.

Description

SignalSilence® Rb siRNA II from Cell Signaling Technology (CST) allows the researcher to specifically inhibit Rb 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 are rigorously tested in-house and have been shown to reduce target protein expression by western analysis.

Directions for Use

CST recommends transfection with 100 nM Rb siRNA II 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.

Background

The retinoblastoma tumor suppressor protein, Rb, regulates cell proliferation by controlling progression through the restriction point within the G1-phase of the cell cycle (1). Rb has three functionally distinct binding domains and interacts with critical regulatory proteins including the E2F family of transcription factors, c-Abl tyrosine kinase, and proteins with a conserved LXCXE motif (2-4). Cell cycle-dependent phosphorylation by a CDK inhibits Rb target binding and allows cell cycle progression (5). Rb inactivation and subsequent cell cycle progression likely requires an initial phosphorylation by cyclin D-CDK4/6 followed by cyclin E-CDK2 phosphorylation (6). Specificity of different CDK/cyclin complexes has been observed in vitro (6-8) and cyclin D1 is required for Ser780 phosphorylation in vivo (9).

  1. Sherr, C.J. (1996) Science 274, 1672-7.
  2. Nevins, J.R. (1992) Science 258, 424-9.
  3. Welch, P.J. and Wang, J.Y. (1993) Cell 75, 779-90.
  4. Hu, Q.J. et al. (1990) EMBO J 9, 1147-55.
  5. Knudsen, E.S. and Wang, J.Y. (1997) Mol Cell Biol 17, 5771-83.
  6. Lundberg, A.S. and Weinberg, R.A. (1998) Mol Cell Biol 18, 753-61.
  7. Connell-Crowley, L. et al. (1997) Mol Biol Cell 8, 287-301.
  8. Kitagawa, M. et al. (1996) EMBO J 15, 7060-9.
  9. Geng, Y. et al. (2001) Proc Natl Acad Sci USA 98, 194-9.

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.

This product is intended for research purposes only. The product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

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