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siRNA knockdowns, immunoprecipitation and western blotting

CL C M Lucas
MM M Milani
MB M Butterworth
NC N Carmell
LS L J Scott
RC R E Clark
GC G M Cohen
SV S Varadarajan
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Cells were reverse-transfected with 10 nm of BAK (s1880 and s1881), BAX (s1888 and s1889), BIM (s195011), PUMA (pool of siRNAs), BMF (pool of siRNAs), BIK (s1989 and s1990), HRK (s194952), BCL-XL (s1920), MCL-1 (s8583), BCL-w (s1924), BFL-1 (pool of siRNAs) from Life Technologies (Paisley, UK), BID (SI02654568), NOXA (SI00129430), BAD (SI00299348), BCL-2 (S100299411) from Qiagen (Manchester, UK) using Interferin (Polyplus Transfection, NY, USA), according to the manufacturer's protocol and processed 48 h after transfection. Immunoprecipitation and western blotting were carried out according to the standard protocols.26

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