Proteomics

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RNA binding protein hnRNPF deficiency drives malignant transformation


ABSTRACT: 1× 10e7 32D cells were lysed in 1 ml of hypotonic lysis buffer CEB-A (Applygen, Cat# 1200). Cells were disrupted by vortexing vigorously for 30 seconds, followed by incubation on ice for 15 minutes. During this period, samples were vortexed for 15 seconds every 5 minutes. Lysates were then centrifuged at 12,000g at 4°C for 5 minutes. The supernatant, containing the cytoplasmic protein fraction, was carefully transferred to a new tube without disturbing the pellet. The cytoplasmic lysates were incubated overnight at 4°C with either an HnRNPF antibody or IgG as a control. Protein G magnetic beads were added, and incubation continued for 2 hours at 4°C. After incubation, the beads were washed four times with wash buffer (50 mM Tris-HCl, 140 mM NaCl, 0.1% NP-40, 1 mM EDTA, 10% Glycerol), and the target proteins were eluted. The eluted proteins were resolved on a gradient gel and visualized by silver staining. Specific bands corresponding to both the control and experimental groups were excised and sent to Shanghai Applied Protein Technology (Shanghai, China) for mass spectrometry analysis to identify proteins interacting with HNRNPF in the cytoplasm.

ORGANISM(S): Mus Musculus

SUBMITTER: Tao Cheng  

PROVIDER: PXD062334 | iProX | Fri Mar 28 00:00:00 GMT 2025

REPOSITORIES: iProX

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