Cells were lysed in RIPA buffer containing protease inhibitor cocktail (Roche) and 1 mM phenylmethylsulfonyl fluoride (PMSF). Equal amount of protein lysate was resolved on a 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to a nitrocellulose membrane (Millipore) using fast Turbo Transfer (Bio-rad). After blocking in 5% skimmed milk for 1 h, the appropriate primary antibody was added: anti-ADAR2 (RED1) (1:1000, ab64830, Abcam), anti-SRp30c(SRSF9) (1:1000, sc-134036, Santa Cruz), anti-FLAG (1:2000, Clone M2, Sigma), anti-cMyc (1:1000, sc-789, Santa Cruz), anti-V5 (1:2000, ab9116, Abcam) or anti-β-actin (1:1000, clone C4, sc-47778, Santa Cruz). Primary antibodies were incubated overnight in the cold room. After washing with phosphate-buffered saline (PBS)/0.2% Tween-20 (PBST), a secondary antibody, namely horse-radish peroxidase (HRP)-conjugated anti-rabbit IgG (1:5000, NA934V, Amersham) or HRP-conjugated anti-mouse IgG (1:5000, NA931V, Amersham), was added for 1 h. After washing with PBST, signals were detected using the WesternBright Sirius Chemiluminescent HRP Substrate (Advansta).
Sc 134036
Manufactured by Santa Cruz Biotechnology
Sourced in United States
Sc-134036 is a laboratory product manufactured by Santa Cruz Biotechnology. It is a piece of equipment designed for use in research and scientific applications. The core function of this product is to provide a specific capability or tool for conducting experiments or analyses. However, a detailed description of the product's features, specifications, or intended use cannot be provided while maintaining an unbiased and factual approach.
Automatically generated - may contain errors
Lab products found in correlation
Ab64830, by Abcam (1 mentions)
Nitrocellulose membrane, by Merck Group (1 mentions)
Ab9116, by Abcam (1 mentions)
Sc 789, by Santa Cruz Biotechnology (1 mentions)
Sc 47778, by Santa Cruz Biotechnology (1 mentions)
Clone c4, by Santa Cruz Biotechnology (1 mentions)
Clone m2, by Merck Group (1 mentions)
Westernbright sirius chemiluminescent hrp substrate, by Advansta (1 mentions)
Na934v, by Cytiva (1 mentions)
Na931v, by Cytiva (1 mentions)
Protease inhibitor cocktail, by Roche (1 mentions)
2 protocols using sc 134036
1
Western Blot Analysis of Protein Expression
2
Biotinylated RNA Pulldown Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
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RNA pulldown analysis was performed, as described previously [25 (link)]. Briefly, streptavidin agarose was used to covalently link 5′-end biotin-labeled RNA by incubating with buffer D (20 mM Tris-Cl pH 7.5, 150 mM KCl, 0.2 mM EDTA, 10% glycerol, 0.5 mM DTT, 0.5 mM PMSF) at 4 °C for 1h. RNA-linked streptavidin beads were incubated with cell lysates for 4 h at 4 °C and then washed with buffer D five times. Beads were loaded onto 10% SDS-PAGE gel and subjected to immunoblotting analysis using anti-Tra2β (Santa Cruz, sc-166829, Dallas, TX, USA) and anti-SRSF9 (Santa Cruz, sc-134036) antibodies. Biotin-labeled RNA sequences are shown in Table S1 .
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