GSM6935174: H3K27ac_ChIPseq_WT2; Mus musculus; ChIP-Seq
Following sorting, ChIP was performed using the ChIP-IT High Sensitivity Kit (Active Motif, Cat. No. 53040). The procedure followed the manufacturer recommended protocol with specific modifications: (i) the nuclear pellet of 10M cells was lysed in 500 μl ChIP buffer and the chromatin was sheared to an average of 200 to 600 base pairs using the Sonicator Q125 (QSonica, Newtown, CT, USA); (ii) the immunoprecipitation was performed with 4 μL of anti-acetyl-H3 (Lys27) (D5E4) (Cell Signaling Technology, Cat. No. 8173). Sequencing libraries were prepared using the Next Gen DNA Library kit (Active Motif, Cat. No. 53216) and Next Gen Indexing Kit (Active Motif, Cat. No. 53264) for Illumina. Libraries were purified with the Magnetic SPRI beads (Beckman Coulter #B23317) and amplified with 9 PCR cycles (Mastercycler Nexus, Eppendorf). Quality of amplified libraries was validated using ChIP-IT qPCR Analysis Kit (Active Motif, Cat. No. 53029) and libraries were sequenced as PE150 on Illumina NovaSeq 6000.