GSM2199457: JS.mycn-high; Homo sapiens; ChIP-Seq
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody. Chromatin immunoprecipitated DNA was prepared for sequencing following a modified version of Illumina's Genomic DNA protocol. Sheared DNA fragments undergo end repair by T4 DNA polymerase to fill in 5′ overhangs, Klenow polymerase to remove 3′ overhangs, and T4PNK to phosphorylate the 5′-OH. For Illumina adapter ligation, a single adenine nucleotide overhang must be added to the polished DNA. A 1:30 dilution of the Adaptor Oligo Mix (20) was used in the ligation step. The resulting adapted constructs are amplified by 18 cycles of PCR enriching the DNA fragments with adapters flanking each end. The amplified product was quantified using a spectrophotometer at 260 nm (Nanodrop) then a portion of final sample is size separated on a 6% TBE (Invitrogen) to verify correct target range amplification. The validated DNA library is submitted at a 10 nmol/L concentration for sequencing. All protocols for Illumina library sequence preparation, sequencing, and quality control are provided by Illumina.
GEO
GSM2199457