title: GSM9120528: iPSC-CM_IndA_Untr_Input; Homo sapiens; ChIP-Seq
library_construction_protocol: Chromatin immunoprecipitation (ChIP) was performed to map TOP2B binding sites in untreated iPSC-derived cardiomyocytes from individual UCSD143i-87-1. Approximately 40 million cells were fixed in 1% formaldehyde, quenched with glycine, and flash-frozen. Chromatin was extracted, sequentially lysed, and sonicated using a Covaris system to generate 250–400 bp fragments. Immunoprecipitation was performed overnight at 4 °C using anti-TOP2B antibody (Abcam ab72334) pre-bound to Dynabeads Protein G. After stringent washes, chromatin was eluted and crosslinks reversed. DNA was purified by phenol-chloroform extraction and ethanol precipitation, then resuspended for library preparation. ChIP and Input DNA libraries were constructed using the TruSeq ChIP Sample Preparation Kit (Illumina, IP-202-1012) following manufacturer instructions with minor modifications. Starting with all the ChIP DNA and 50ng of  input DNA, end repair and A-tailing were performed prior to ligation of indexed adapters (ACAGTG for ChIP, GCCAAT for Input). Adapter-ligated DNA was PCR-amplified for 18 cycles and purified using AMPure XP beads. Libraries were size-selected (250–400 bp) by gel excision, then purified with the MinElute Gel Extraction Kit (Qiagen). Library concentration and size distribution were assessed by Qubit and Agilent Bioanalyzer. Final libraries were sequenced as 40 bp paired-end reads: ChIP library on the Illumina NextSeq 550 (generated 40,380,049 reads) and Input library on the Illumina MiniSeq (20,968,544 reads).