Hmgn2

The following primary rabbit antibodies were purchased from Abcam: anti-H1.2, ab17677; H1.5, ab18208; HMGB2, ab124670; Ki67, ab15580; H3S10p, ab5176; CTCF, ab128873; SMC2, ab10412; RAD21, ab154769; H1x, ab31972. From Cell Signaling: anti-HMGN1, #5692; HMGN2, #9437. From Sigma-Aldrich: anti-H1.4, H7665. From Millipore: anti-H2A, #07-146. The following mouse antibodies were purchased from Abcam: anti-H3, ab24834; H4, ab31830; H2B, ab52584. From Active Motif: anti-H3, #61475. mAb PL2-6 was a gift from M. Monestier (Temple Univ.), see previous use [26,27]. Secondary antibodies included Invitrogen Alexa 568 goat anti-rabbit IgG, Sigma-Aldrich Atto 488 goat anti-rabbit IgG and Atto 488 goat anti-mouse IgG.

Tissue sample (100 µg) was added to the RIPA buffer (Thermo, Waltham, MA, USA) for homogenization and then subjected to centrifugation to obtain the supernatant. Protein concentration was quantified using the BCA assay kit (Thermo, Waltham, Massachusetts). The tissue lysates were separated using 10% SDS–PAGE gel for 90 min and transferred to polyvinylidenefluoride membranes (Roche, Basel, Switzerland) for 2 h. After blocking with 5% BSA in Tris-buffered saline for 1 h. Then, the blots were incubated with primary antibodies for FoxA1 (Cell signaling, Beverly, MA, USA), HMGN1 (Cell signaling, Beverly, MA, USA), HMGN2 (Cell signaling, Beverly, MA, USA), HMGN3 (Abcam, Cambridge, UK), and TSBP1 (Abcam, Cambridge, UK) overnight, after which they were reacted with an HRP-conjugated secondary antibody for 2 h. An enhanced chemiluminescence prime western blotting detection reagent (GE Healthcare, Chalfont St. Giles, UK) was used to visualize the protein bands. The density of respective bands was analyzed with the iBright 1500 (Thermo, Waltham, MA, USA).