Anti cbp sc 33000

Liver tissues or cell pellets were lysed in hypotonic buffer (5 mM Tris-HCl, 1 mM MgCl₂, 3 mM CaCl₂, 8% Sucrose). After centrifuge, pellets were washed once with hypotonic buffer and re-suspended in RIPA buffer (5 mM Tris-HCl, 150 mM NaCl, 0.1% SDS, 0.05% deoxycholic acid, 10% Glycerol) prior to sonication for 5 s. Nuclear fractions were then collected after centrifugation at 13,000 rpm × 10 min. FLAG-M2 beads or streptavidin beads were added into nuclear fractions to capture FLAG-CRY1 or CBP-CRY1. The protocol for detecting ubiquitination was reported [20 (link)] with minor modifications. anti-ChREBP was used to pull down poly-ubiquitinated ChREBPα conjugates. Western blot analysis was performed using the following primary antibodies: anti-DDB1 (Abcam ab9194), anti-CRY1 (sc-101006), anti-GAPDH (sc-25778), anti-Lamin A/C (sc-20681), anti-CBP (sc-33000) (Santa Cruz biotechnology), anti-ChREBP (Novus NB400–135), anti-ubiquitin (Sigma U5379), and anti-β-tubulin (T5201) (Sigma-Aldrich).