Cytoplasmic and Nuclear Protein Extraction

Cytoplasmic and nuclear proteins were extracted using the Nuclear and Cytoplasmic Protein Extraction Kit (Beyotime). Briefly, cells were harvested by centrifugation and resuspended in cytoplasmic extraction agent A. The solution was vortexed and incubated on ice for 10 min. Cytoplasmic extraction agent B was then added to the cell pellet and the pellet was vortexed and incubated on ice. The supernatant was collected as the cytoplasmic extract. The insoluble fraction was suspended in nuclear extraction agent. After ultrasonication for 30 min, the mixture was centrifuged and the supernatant was collected as the nuclear extract.