title: GSM4176114: Mouse HSC H3K27ac 1 week recovery; Mus musculus; ChIP-Seq
library_construction_protocol: For Chip-seq Lysates were clarified from sonicated nuclei and protein-DNA complexes were isolated with antibody. Alternatively, DNA was phenol/chloroform-extracted from MNase-treated nuclei, and 134-154 bp long fragments were extractred from a 2% agarose gel. For RNA-seq Total RNA was isolated using Purelink RNA Mini Kit (Life Technologies) For Chip-seq Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3' end. After adapter ligation, ChIP DNA was PCR-amplified with Illumina primers for 12-16 cycles and library fragments of 200-300 bp (ChIP) (insert plus adaptor and PCR primer sequences) were isolated from a 2% agarose gel. For MNase-Seq, size-selected mononucleosome-derived DNA fragments were ligated to modified adapters that incorporate both the adapter sequence as well as the primer landing sites for the flow cell-conjugated primers. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer or Genome Analyzer II following the manufacturer's protocols. For RNA-seq Illumina Stranded Total RNA (Illumina 20020597)
identifiers.namespace: GEO
identifiers.id: GSM4176114