title: GSM9166780: Huvec cells,WT,low glucose (5 mM),12 hours; Homo sapiens; ChIP-Seq
library_name: GSM9166780
library_construction_protocol: Nuclei were isolated from fresh or lightly fixed cells and incubated with a primary antibody specific to the target protein. After washing, cells were incubated with a secondary antibody conjugated to a pA-Tn5 transposase fusion protein. The tethered Tn5 transposase was activated by magnesium to simultaneously cleave and tag DNA at antibody-bound sites. Tagged fragments were then extracted for library preparation. CUT&Tag libraries were constructed using the CUT&Tag_Novogene Kit following the manufacturer's protocol. Briefly, Concanavalin A (ConA) magnetic beads were used to immobilize cells or nuclei. Membrane permeability was enhanced by digitonin treatment. Target protein binding was achieved via a specific primary antibody, followed by incubation with a secondary antibody. Protein A-Tn5 transposase was then recruited to the target region in a directed manner, enabling in situ tagmentation and simultaneous addition of sequencing adapters. The tagmented DNA fragments were PCR amplified to generate sequencing-ready libraries. Post-amplification libraries were purified using AMPure XP beads (Beckman Coulter) and evaluated for quality using the Agilent Bioanalyzer 2100 system.