title: GSM8098649: Endogenously tagged ZFP661-3HA mice, E16.5 forebrain (replicate 1), ChIP input; Mus musculus; ChIP-Seq
library_construction_protocol: Embryonic forebrains were isolated and dissociated into a single-cell suspension using the Papain Dissociation System (Worthington LK003150) by incubating at 37°C for 45 min. The cell suspension was passed through cellTrics Cell Strainers (20 μm) to remove cell debris and clumps. Subsequently, the suspension underwent discontinuous density gradient centrifugation using the Papain Dissociation System at 70 g for 6 min to further remove any remaining debris and membrane fragments. Then, cells were crosslinked with 1% formaldehyde for 10 min and quenched with 0.4M glycine for 5 min. ChIP assays were performed according to the ENCODE protocol with modifications. Eight E16.5 forebrains per ChIP were used for ZFP661-3HA (anti-HA: Abcam, ab9110) and CTCF (anti-CTCF: Millipore, 07-729). 5 μg of antibodies were used for each ChIP. ChIP DNA was purified, and 10 μl of ChIP DNA was used as input to construct the ChIP library using the ThruPLEX DNA-Seq Kit (Takara, R400676). The ChIP inputs from the same biological replicate were used to construct the corresponding ChIP input library.