Western blot analysis of extracts from HeLa cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-) or SignalSilence® hnRNP A1 siRNA I (+), using hnRNP A1 (D21H11) Rabbit mAb #8443 and α-Tubulin (11H10) Rabbit mAb #2125. The hnRNP A1 (D21H11) Rabbit mAb confirms silencing of hnRNP A1 expression, while the α-Tubulin (11H10) Rabbit mAb is used as a loading control.
CST recommends transfection with 100 nM SignalSilence® hnRNP A1 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.
SignalSilence® siRNA is supplied in RNAse-free water. Aliquot and store at -20ºC.
SignalSilence® hnRNP A1 siRNA I from Cell Signaling Technology (CST) allows the researcher to specifically inhibit hnRNP A1 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.
Heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) is a member of the hnRNP A/B family of related RNA binding proteins that bind pre-mRNA and are involved in the processing, metabolism, and transport of nuclear pre-mRNA transcripts (1). hnRNP A1 regulates the alternative splicing of c-Src and c-H-Ras (2,3) and modifies initiation of translation of the fibroblast growth factor 2 mRNA (4). hnRNP A1 expression level is elevated in many cancers; knockdown of hnRNP A1 leads to apoptosis in various cancer cells (5). Although predominantly nuclear, hnRNP A1 is continually transported from the nucleus to the cytoplasm where it disassociates from mRNA and is rapidly re-imported into the nucleus (6,7). hnRNP A1 binds to cis-acting repressive sequences (CRS) of HIV-1 to influence HIV-1 production (8,9). HIV-1 enhances hnRNP A1 expression and promotes the relocalization of hnRNP A1 to the cytoplasm (10).
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