embryonic stem cell line derived from mouse inner-cell masses
stain
12910la
chip antibody
HP1gamma (Millipore, MAB3450, lot 2021184)
trasfection
tranfected with siHP1g for 48h
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody. Libraries were prepared according toThruPLEX® DNA-seq Kit Instruction Manual (CAT. NO. R400406). 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 DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols.