title: GSM1944093: mutant 1 H3K4me2 ChIP-Seq; Mus musculus; ChIP-Seq
library_construction_protocol: ChIP-Seq Library Prep: Immunoprecipitated complexes were washed and isolated.  Crosslinks were reversed overnight (65C, 0.54M NaCl).  DNA was then isolated and treated with T4 DNA polymerase, E. coli Pol I large (Klenow) fragment, and T4 polynucleotide kinase to ensure all dsDNA fragments are blunt-ended with 5' phosphates.  DNA then repurified and treated with Klenow 3' -> 5' exo minus and dATP to adenylate the 3' end of each fragment.  DNA was repurified and ligated to Illumina TruSeq indexed adapters containing a T-overhang using T4 DNA ligase.  DNA was repurified and PCR amplified using primers complimentary to the Illumina TruSeq adapter sequence.  Amplified fragments between 200 and 500bp were isolated from an agarose gel and purified.  This DNA was then clustered on an Illumina flow cell and sequenced on an Illumina HiSeq 2500. Both WT and mutant hearts were thawed on ice and pooled into 3 replicates of 30 hearts each. Samples were crosslinked with 1% formaldehyde, lysed with cell lysis and nuclei lysis buffer, and sonicated to shear chromatin into 200-600 bp fragments. Each replicate was further split into input and 3 samples. Chromatin immunoprecipitation was performed with 3 ug of anti-H3K4me1/2/3 antibody (Abcam) overnight with Protein G Dynabead (Life Technologies). After washing, DNA was eluted, reverse-crosslinked and precipitated with AmPure XP magnetic beads (Beckman Coulter).
attributes.tag: GEO Accession
attributes.value: GSM1944093