GSM9206955: Gadd45a knockout mice, Input, Prefrontal cortex, biol rep1; Mus musculus; ChIP-Seq
Tissue fixation was carried out using 1% formaldehyde for 10 minutes at room temperature under a vacuum pump. The crosslinking reaction was terminated by adding 0.125 M glycine and allowing the mixture to settle for 5 minutes. Subsequently, the tissue was collected, frozen in liquid nitrogen, and processed using a tissue lyser. The resulting powder underwent treatment with cell lysis buffer, and the nucleus was isolated by centrifugation at 2000g for 5 minutes. The isolated nucleus was then subjected to nucleus lysis buffer treatment and sonication to fragment the chromatin DNA. Following sonication, 10% of the lysed chromatin was stored as “input,” while 45% was utilized in immunoprecipitation reactions with an anti-TET1 antibody (GeneTex, GTX124207) and labeled as “IP.” Additionally, 45% was incubated with rabbit IgG (Cell Signaling Technology) as a negative control, labeled as “IgG.” Subsequently, DNA extraction from both input and IP samples was performed using the phenol-chloroform method. High-throughput DNA sequencing libraries were prepared using the VAHTS Universal DNA Library Prep Kit for Illumina V3 (Catalog NO. ND607, Vazyme).