alias: GSM3408775
title: GSM3408775: Tip60 KO HSPC H3K27ac; Mus musculus; ChIP-Seq
library_construction_protocol: RNA was isolated using RNeasy mini columns (Qiagen). As for ChIP, briefly, cells were fixed with 1% formaldehyde and then neutralized with glycine at a final concentration of 0.125M. Fixed chromatin were fragmented using a Bioruptor sonicator (Diagenode) to an average size around 300-500 base pairs. Sonicated chromatin was incubated with antibody overnight at 4°C. The antibodies used are Tip60 50, H2A.Z (abcam, ab4174), acH2A.Z (abcam, ab18262), H3K27ac (abcam, ab4729), and H3K27me3 (CST, 9733BC). Protein A and Protein G Dynabeads (Life Technologies) were added to the mixture and incubated for 2 hours at 4°C. Antibody:chromatin complexes were collected with a magnet and washed two times with a solution of 50 mM HEPES-KOH, pH 7.6, 500 mM LiCl, 1 mM EDTA, 1% NP-40 and 0.7% Na-Deoyxcholate. After a TE wash, samples are eluted, RNase and proteinase K treated and purified using a QIAGEN PCR purification kit. A mimunim of 10 ng of DNA was used for library construction using ThruPLEX kits (Rubicon Genomics). RNA-seq libraries were prepared using Illumina ScriptSeq™ v2 RNA-Seq  Library Preparation Kit, according to the manufacturer's instructions (Illumina, San Diego, California, USA). Libraries were validated with an Agilent Bioanalyzer (Agilent Technologies, Palo Alto, CA), diluted and applied to an Illumina flow cell using the Illumina Cluster Station. ChIP-seq libraries were prepared using ThruPLEX® DNA-seq Kit (Rubicon), and multiplexed.
identifiers.namespace: GEO
identifiers.id: GSM3408775
attributes.tag: GEO Accession
attributes.value: GSM3408775