GSM2776413: ACTD; Mus musculus; OTHER
Treated cells (4X15 cm dishes)  were harvested by trypsinization and rinsed twice by PBS. Then the cells were treated by 10ml Buffer A (10mM Hepes, pH 7.9, 10mM KCl, 1.5mM MgCl2, 0.5mM DTT) on ice for 5min followed by 10X Dounce homogenization. Centrifuge the cells at 218g for 5min at 4C. Then resuspend the pellet with 3 ml S1 solution (0.25M sucrose, 10mM MgCl2) and layered over 3ml S2 solution (0.35M sucrose, 0.5mM MgCl2) and centrifuged at 1430g for 5min at 4C. Resuspend the pellet with 3ml of S2 solution and sonicate to break the nuclei. Layered the sonicated sample over 3ml of S3 solution (0.88M Sucrose, 0.5mM MgCl2) and centrifuge at 3000g for 10min at 4 C. Resuspended the nucleoli with 500ul of S2 solution and centrifuged for another 5min. The pellet was resuspended and purified by phenol-chloroform extraction. Nucleolar retrieved DNA-seq were prepared by library preparation modules (New England Biolabs)