Western blot analysis of extracts from C2C12 cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-), SignalSilence® β-Catenin siRNA I (Mouse Specific) #6387 (+) or SignalSilence® β-Catenin siRNA II (Mouse Specific) (+), using β-Catenin (6B3) Rabbit mAb #9582 (upper) or α-Tubulin (11H10) Rabbit mAb #2125 (lower). The β-Catenin (6B3) Rabbit mAb confirms silencing of β-Catenin expression, while the α-Tubulin (11H10) Rabbit mAb is used as a loading control.
CST recommends transfection with 100 nM β-Catenin siRNA II (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.
SignalSilence® siRNA is supplied in RNAse-free water. Aliquot and store at -20ºC.
SignalSilence® β-Catenin siRNA II (Mouse Specific) from Cell Signaling Technology (CST) allows the researcher to specifically inhibit β-catenin 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.
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.
β-Catenin is a key downstream effector in the Wnt signaling pathway (1). It is implicated in two major biological processes in vertebrates: early embryonic development (2) and tumorigenesis (3). CK1 phosphorylates β-catenin at Ser45. This phosphorylation event primes β-catenin for subsequent phosphorylation by GSK-3β (4-6). GSK-3β destabilizes β-catenin by phosphorylating it at Ser33, Ser37, and Thr41 (7). Mutations at these sites result in the stabilization of β-catenin protein levels and have been found in many tumor cell lines (8).
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SignalSilence is a registered trademark of Cell Signaling Technology, Inc.