title: GSM6423445: 12Z_shARID1A_H3.1_rep2_ChIP; Homo sapiens; ChIP-Seq
library_construction_protocol: Cells were treated with 1% formaldehyde in growth media for 10 minutes at ambient temperature. Formaldehyde was quenched by the addition of 0.125 M Glycine and incubation for 5 minutes at room temperature, followed by PBS wash and scraping. 1*10^7 crosslinked cells were used for each ChIP, and ChIP was performed in duplicate. Chromatin from crosslinked cells was fractionated by digestion with micrococcal nuclease using the SimpleChIP Enzymatic Chromatin IP Kit (Cell Signaling) according to the manufacturer protocol, followed by 30 seconds of sonication. ChIP was then performed according to the SimpleChIP Enzymatic Chromatin IP Kit (Cell Signaling) with the addition of 5 mM sodium butyrate to preserve histone acetylation. 1:250 anti-histone H2A.Z (ab4174, abcam) or 1:50 anti-acetyl-histone H4 (06-866, Millipore) was added to each, respective 1.25 mL IP. Crosslinks were reversed with 0.4 mg/mL Proteinase K (ThermoFisher) and 0.2 M NaCl at 65 °C for 2 hours. DNA was purified using the ChIP DNA Clean & Concentrator Kit (Zymo). Libraries for input and IP samples were prepared by the Van Andel Research Institute Genomics Core. 10 ng of material was used for input samples, and the entire precipitated sample was used for IPs. Libraries were generated using the KAPA Hyper Prep Kit (v5.16) (Kapa Biosystems). Prior to PCR amplification, end-repaired and A-tailed DNA fragments were ligated to IDT for Illumina UDI Adapters (IDT DNA Inc.). Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies), QuantiFluor® dsDNA System (Promega) and Kapa Illumina Library Quantification qPCR assays (Kapa Biosystems). Individually indexed libraries were pooled, and 100 bp, single-end sequencing was performed on an Illumina NovaSeq 6000 sequencer using a 100-cycle sequencing kit (Illumina). Each library was sequenced to minimum read depth of 80 million reads per input library and 40 million reads per IP library. Base calling was performed by Illumina NCS v2.0, and NCS output was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v1.9.0.