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2 protocols using microrna 31 mimic

1

TNF-α-mediated Signaling Pathway Analysis

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TNF-α was from R & D Systems. Secondary antibodies and enhanced chemiluminescence reagents were from Amersham. Anti-TRADD and anti-p65 antibodies were from Santa Cruz Biotechnology. Anti-phosphorylated p65 (p-p65) and anti-AcH3 antibodies were from Cell Signaling, and anti-GAPDH and anti-H3K4Me3 antibodies were from Abcam. Real time qRT primers and Taqman qRT-PCR reagents were from Applied Biosystems. Sybr green qRT-PCR reagents were from Qiagen. MicroRNA-31 mimic, miR-31 antagomiR and negative control (−CT) miR were purchased from Ambion/Life Technologies. MicroRNAs were transfected using Lipofectamine RNAiMax Transfection reagent (Life Technologies).
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2

Molecular Profiling of Pluripotent Stem Cells

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IL-1β and TNF-α were from R & D Systems. The secondary peroxidase-conjugated antibodies and enhanced chemiluminescence reagents were purchased from Amersham (Arlington Heights, IL). Anti-KLF4, anti-Lin28, anti-Nestin, anti-Oct4 and anti-SOX2 antibodies were from Santa Cruz. Anti-Olig2 antibodies were purchased from R & D Systems. Anti-GFAP antibodies were from Pharmingen, and anti-Brn2 (POU3F2) and anti-GAPDH antibodies were from Abcam. Taqman qRT-PCR reagents were from Applied Biosystems (Beverly, MA). Sybr green qRT-PCR reagents were from Qiagen (Valencia, CA). All other primers were purchased from Integrated DNA Technologies (Iowa City, IA). MicroRNA-31 mimic, miR-31 AntagomiR and negative control (CT) miR were purchased from Ambion/Life Technologies (Austin, TX). MicroRNAs and shRNA molecules were transfected using Lipofectamine RNAiMax Transfection reagent (Life Technologies, Grand Island, NY). Control and Lin28 specific shRNA were purchased from Dharmacon.
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