title: GSM8657605: DIvA cell, IDH mutant, non treated, Knock down (si2), 4C-seq, DSB2, Rep 1; Homo sapiens; OTHER
library_name: GSM8657605
library_construction_protocol: ChIP-seq was performed using ChIP-IT High Sensitivity (Active Motif, 53040) according to the manufacturer's instructions. Cells were fixed with 1% formaldehyde for 10 minutes. After quenching and cell lysis, sonication was performed with Covaris S220 at a power of 140 W for 20 min. For 4C-seq, The 4C–seq experiments were performed as described 24 with minor modifications. In brief, fifteen million DIvA cells were cross-linked with 2% formaldehyde for 10 min at room temperature, lysed and digested with DpnI (100 U, NEB). Digested DNA was then ligated with 50 U T4 DNA ligase (Promega), purified and digested with NlaIII overnight (NEB). After a second ligation step, DNA was purified before proceeding to library preparation. ChIP library construction was performed with NEBNext® Ultra™ II DNA Library Prep Kit for Illumina. Briefly, the steps include End Repair, dA-Tailing, Adaptor Ligation, ligation products purification and PCR process. 20 ng DNA was used for the starting material. For 4C–seq library preparation, eight hundred nanograms of 4C–seq template were amplified using two-step PCR reactions with inverse primers (PAGE-purified) and primers with Illumina adaptor sequences and a unique index for each condition. Libraries were purified with the QIAquick PCR Purification Kit (Qiagen), pooled and subjected to 150-bp single-end sequencing on the Illumina NextSeq2000 platform at the NCI/CCR Next Generation Sequencing Core Facility.
identifiers.namespace: GEO
identifiers.id: GSM8657605_r1