title: GSM4198706: R-TEn-p1-Ebox_Mut_DP_CTCF; Mus musculus; ChIP-Seq
library_construction_protocol: ATAC-seq : ATAC-seq was performed as described (Buenrostro et al., 2013) with modifications. Cells were lysed, then treated with Tn5 transposase (Nextera DNA Sample Preparation Kit). After the treatment, the tagmentated DNA was cleaned up and purified with DNA Clean & Concentrator kit (Zymo Research).  The purified DNA were amplified with determined cycles of PCR based on the amplification curve, then were size-selected using Ampure XP beads. Hi-C : in situ Hi-C was performed as described (Rao et al., 2014).  DP and cultured proB cells from control and deletion mutant mice were fixed with 1% formaldehyde/FACS buffer for 10 minutes at room temperature. Cells were lysed and chromatin was digested overnight using MboI. The fragment overhangs generated by the restriction enzyme were filled in with biotin-14-dATP by Klenow fragment, then proximity ligation was performed.  Biotinylated DNA was sonicated to 300-500 bp on a Covaris M220. The biotinylated DNA was captured on Dynabeads MyOne Streptavidin T1 beads. After washing the beads, ends of sheared DNA were repaired and added A-tail according to the protocol. NEBNext Multiplex Oligos for Illumina (NEB) were ligated and the beads were washed.  Libraries were amplified by PCR and size selected using ProNex (Promega). As to CD4SP cells, Tethered Conformation Capture (TCC) was performed as described (Kalhor et al. 2012).  CD4SP cells were fixed with 1% formaldehyde/FACS buffer for 10 minutes at room temperature. Cells were lysed and chromatin was digested overnight using MboI.  ChIP-seq : Cells were fixed in 1% formaldehyde/PBS . The reactions were quenched by 1.5M glycine and cells were washed with PBS and pelleted. Pelleted cells were lysed and chromatin was sonicated by Covaris M220.  Diluted sheared chromatin was immunoprecipitated with each antibody which is pre-bound to Dynabead Protein G magnetic microbeads overnight at 4 °C. Samples were washed, eluted and reverse cross-linked  65°C overnight. Samples were next treated with RNase A and proteinase K and purified using DNA Clean & Concentrator kit (Zymo Research). Samples were ligated to NEBNext Multiplex Oligos for Illumina (NEB) and multiplex amplified by PCR. Amplified ChIP DNA was selected by size.