Anti gfp

The following antibodies were purchased from Cell Signaling Technology: anti-IRS-1 (2382), anti-pAKT S473 (9271), anti-p85 (4292), anti-IGF-1Rβ (9750), anti-PDK1 (13037). Other antibodies were from the following commercial sources: anti-VAPB (Sigma-Aldrich, HPA013144), anti-NOGOA (Bio-Rad, AHP1799), anti-BAP31 (Santa Cruz Biotechnology, sc-48766), anti-PERK (abcam, ab229912), anti-pPERK (Affinity Biosciences, DF7576), anti-AKT (HUABIO, ET1609-47), anti-Actin (HUABIO, M1210-2), anti-Tubulin (HUABIO, M1305-2), anti-GFP (HUABIO, ET1607-31), anti-FLAG (YEASEN, 30503ES60), anti-HA (Invitrogen, PA1-985) and anti-mCherry (ABclonal, AE002).

The mycelia (0.1 g) cultured in liquid CM for 2 days were collected at 28 °C, and the total protein solutions were extracted with lysis buffer (50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1 mM EDTA, 1% Triton 100, and 1 × protein inhibitor) at 12000 rpm for 10 min at 4 °C. For detection of Porin (mitochondria outer-membrane protein), total proteins were extracted with 10% SDS solution. The resultant protein solution was resolved by 8–15% SDS polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to a PVDF membrane, followed by incubation with primary antibodies anti-GFP (Huabio, ET1607-31, Hangzhou, China) or anti-porin antibody (GenScript, A01419, Nanjing, China). The protein GAPDH served as a loading control. Western blots were detected using an ECL chemiluminescent kit (Biorad GS-710, Hercules, CA, USA), and the relative intensity of blots was quantified by Image J software (version 1.37).