Cell Signaling Technology

Product Pathways - Cytoskeletal Signaling

SignalSilence® Sec24B siRNA I #12229

Applications Reactivity
Transfection H (Mk)

Reactivity Key:  H=Human  Mk=Monkey
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 (Unconjugated) #6568 (-), SignalSilence® Sec24B siRNA I (+), or SignalSilence® Sec24B siRNA II #12333 (+), using Sec24B Antibody #7427 (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower). The Sec24B Antibody confirms silencing of Sec24B expression, while the β-Actin (D6A8) Rabbit mAb is used as a loading control.

Description

SignalSilence® Sec24B siRNA I from Cell Signaling Technology (CST) allows the researcher to specifically inhibit Sec24B 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® Sec24B 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

Coat Protein Complex II (COPII) is composed of five cytosolic proteins: Sec23/24 complex, Sec13/31 complex, and Sar1. COPII coat is located at the ER/Golgi interface and is involved in transport of newly synthesized proteins from the ER to the Golgi apparatus (1). COPII formation is initiated through the binding of the activated G protein, Sar1, to the Sec23/24 complex, thereby forming a prebudding complex that directly binds target molecules (1-3). The prebudding complex further recruits Sec13/31 to form mature COPII coat (4,5). The Sec24 subunit of COPII coat is thought to play a critical role in cargo selection (2,6). It binds directly to cargo proteins at the ER and brings them to COPII vesicles through interaction with Sec23. There are four Sec24 isoforms in human cells: Sec24A, Sec24B, Sec24C, and Sec24D (7). In mice, mutations in Sec24B have been linked to developmental defects (8,9).

  1. Aridor, M. et al. (1998) J Cell Biol 141, 61-70.
  2. Miller, E.A. et al. (2003) Cell 114, 497-509.
  3. Mossessova, E. et al. (2003) Cell 114, 483-95.
  4. Barlowe, C. et al. (1994) Cell 77, 895-907.
  5. Bi, X. et al. (2007) Dev Cell 13, 635-45.
  6. Miller, E. et al. (2002) EMBO J 21, 6105-13.
  7. Tang, B.L. et al. (1999) Biochem Biophys Res Commun 258, 679-84.
  8. Merte, J. et al. (2010) Nat Cell Biol 12, 41-6; sup pp 1-8.
  9. Wansleeben, C. et al. (2010) Development 137, 1067-73.

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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