GSM9076993: HRMEC, NG2, PES1 (ChIP); Homo sapiens; ChIP-Seq
Add final concentration of 1% Formaldehyde solution for Cross-Linking of DNA and Protein. Then stoped the cross-linking reaction by adding final concentration of 125mM glycine. Extract chromatin from cross-linked cells by adding Lysis Buffer.Sonicate chromatin DNA to fragments ranging from 100 to 500 bp. The sequencing libraries of immunoprecipitated chromatin DNA were generated with varied index label by NEBNext® UltraTM DNA Library Prep Kit for Illumina (NEB, USA, cat. E7645L). Add End Repair Reaction Buffer (10X)和 End Prep Enzyme Mix for end repair by the following condition: 20°C for 30min, 65°C for 30min. And then NEBNext Adaptor and Blunt/TA Ligase Master Mix was added to ligate adaptor at 20°C for 15min. USER Enzyme was added to remove base U of adaptor at 37°C for 5min. After purification with beads, products are amplified with PCR by the following conditions: initial denaturation at 98℃ for 10 sec; denaturation at 98℃ for 10 sec, annealing at 65℃ for 30 sec, and extension at 72℃ for 30 sec. The library size was selected and then purified with AMPure XP Beads.