14519 - Cell Signaling Technology

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Keywords

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Applications: W—Western IP—Immunoprecipitation IHC—Immunohistochemistry ChIP—Chromatin Immunoprecipitation IF—Immunofluorescence F—Flow cytometry E-P—ELISA-Peptide Species Cross-Reactivity: H—human M—mouse R—rat Hm—hamster
Mk—monkey Mi—mink C—chicken Dm—D. melanogaster X—Xenopus Z—zebrafish B—bovine Dg—dog Pg—pig Sc—S. cerevisiae Ce—C. elegans Hr—Horse All—all species expected Species enclosed in parentheses are predicted to react based on 100% homology.
For Research Use Only. Not For Use In Diagnostic Procedures.
www.cellsignal.com
Support: 877-678-TECH (8324)
info@cellsignal.com
Orders: 877-616-CELL (2355)
orders@cellsignal.com
Thank you for your recent purchase. If you would like
to provide a review visit www.cellsignal.com/comments.
SignalSilence® RAP80 siRNA I
New 10/14
Species Cross-Reactivity: H
Description: SignalSilence® RAP80 siRNA I from Cell Signaling Technology (CST) allows the researcher to specifically
inhibit RAP80 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.
Background: The breast cancer type 1 susceptibility protein
(BRCA1) is an E3 ubiquitin ligase that functions in the maintenance of genome stability through regulation of the DNA damage
response and DNA repair. BRCA1 protein forms at least three
distinct complexes (BRCA1 A, B, and C) with other DNA repair
proteins, and these interactions are vital for regulation of BRCA1
protein function. The BRCA1-RAP80 complex (BRCA1 A complex) includes RAP80, BRCC36, BRE, Abraxas, and NBA1 and
functions in G2/M phase checkpoint control (reviewed in 1,2).
The ubiquitously expressed receptor-associated protein 80
(RAP80, UIMC1) is required for recruitment and stability of the
BRCA1 A complex at sites of DNA damage (3). Research studies
indicate that the absence of RAP80 in cells results in increased
sensitivity to the topoisomerase II inhibitor etoposide (4). In the
absence of functional RAP80, BRCA1 A complex function is suppressed and cells become more sensitive to DNA damage-induced
genome instability (5,6). Phosphorylation of RAP80 by CDK1/
Cyclin B at Ser177 regulates RAP80 function at the mitotic checkpoint (7). A naturally occurring in-frame deletion mutant within
RAP80 likely alters RAP80 protein-protein interactions and is
associated with an increase in chromosomal abnormalities (8,9).
Directions for Use: CST recommends transfection with 100
nM RAP80 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.
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.
Storage: RAP80 siRNA I is supplied in RNAse-free water.
Aliquot and store at -20ºC.
For product specific protocols and a complete listing
of recommended companion products please see the
product web page at www.cellsignal.com
Background References:
(1) Ohta, T. et al. (2011) FEBS Lett 585, 2836-44.
(2) Huen, M.S. et al. (2010) Nat Rev Mol Cell Biol 11, 138-48.
(3) Wu, J. et al. (2012) J Biol Chem 287, 22919-26.
(4) Iijima, J. et al. (2010) Cancer Res 70, 8467-74.
(5) Bian, C. et al. (2012) PLoS One 7, e40406.
(6) Yin, Z. et al. (2012) Cancer Res 72, 5080-90.
(7) Cho, H.J. et al. (2013) J Biol Chem 288, 3768-76.
(8) Nikkilä, J. et al. (2009) Oncogene 28, 1843-52.
(9) Anamika et al. (2014) J Biol Chem 289, 12852-62.
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XP®, SignalSilence® and Cell Signaling Technology® are trademarks of Cell Signaling Technology, Inc.
© 2014 Cell Signaling Technology, Inc.
St or e at -2

C
Western blot analysis of extracts from 293T cells, transfected with
100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-) or
SignalSilence® RAP80 siRNA I (+), using RAP80 (D1T6Q) Rabbit
mAb #14466 (upper) and α-Actinin (D6F6) XP® Rabbit mAb #6487
(lower). The RAP80 (D1T6Q) Rabbit mAb confirms silencing of
RAP80 expression, while the α-Actinin (D6F6) XP® Rabbit mAb is
used as a loading control.
kDa
RAP80
α-Actinin
RAP80 siRNA
140
100
80
60
50
40
140
100
80
60
50
40
– +
UniProt ID #Q96RL1
Entrez-Gene ID #51720

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