Real-Time PCR and Immunoblot Analysis

Real-time PCR analysis was performed with primers and probe sets from Applied Biosystems, as described (Liu et al., 2009 (link)). Immunoblots were performed as described previously (Liu et al., 2010 (link)), using the primary antibodies anti-CD115, anti-phosphorylated (T37/46) 4E-BP1 (236B4), anti-phosphorylated (S235/236) S6 (DX57.2.2E), anti-phosphorylated ERK1/2 (D13.14.4E), anti-phosphorylated (S473) AKT (D9E), anti-S6 (5G10), anti-4E-BP-1 (53H11), anti-ERK1/2 (137F5), anti Akt (polyclonal; all from Cell Signaling Technology), anti-SREBP2 (ab30682; Abcam), and anti-β-actin (AC-74; Sigma-Aldrich) and the secondary horseradish peroxidase–conjugated antibodies anti–rabbit IgG (W401B) and anti–mouse IgG (W402B; both from Promega).

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Karmaus P.W., Herrada A.A., Guy C., Neale G., Dhungana Y., Long L., Vogel P., Avila J., Clish C.B, & Chi H. (2017). Critical roles of mTORC1 signaling and metabolic reprogramming for M-CSF–mediated myelopoiesis. The Journal of Experimental Medicine, 214(9), 2629-2647.

Publication 2017

anti-SREBP2 (ab30682; anti-β-actin (AC-74; anti–rabbit IgG (W401B) anti–mouse IgG (W402B;

4e bp1 Actin Anti igg Antibodies Erk1 Horseradish peroxidase Immunoblots Mouse Primers Promega Rabbit Real time pcr analysis

Corresponding Organization : St. Jude Children's Research Hospital

Other organizations : Broad Institute, Harvard University

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Variable analysis

independent variables

Primers and probe sets from Applied Biosystems

dependent variables

Real-time PCR analysis
Immunoblot analysis of CD115, phosphorylated (T37/46) 4E-BP1, phosphorylated (S235/236) S6, phosphorylated ERK1/2, phosphorylated (S473) AKT, S6, 4E-BP-1, ERK1/2, AKT, SREBP2, and β-actin

control variables

Methods described previously in Liu et al., 2009 and Liu et al., 2010

controls

Positive control: Not specified
Negative control: Not specified

Annotations

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