GSM2552052: UMCG-TT-18 Responder; Homo sapiens; MBD-Seq
DNA extracted using Qiagen Blood and Tissue Kit according to manufacturer's instructions. Libraries were prepared using standard multiplexed paired-end ChIP-seq protocol (Illumina). In summary, 500 ng of DNA per sample was fragmented by ultrasound mediated DNA shearing (Covaris S2, Covaris, Woburn, Massachusetts, USA) set to obtain fragments with an average length of 200bp. We employed the MethylCap kit (Diagenode AF-100-0048, Liège, Belgium) to obtain methylated DNA fragments using the highest salt concentration single elution step option. This was followed by the library preparation for the enriched DNA, which was performed with a modified 'multiplexed paired-end ChIP protocol' (Illumina, San Diego, California, USA). Therefore, the NEBNext® DNA Sample Prep Master Mix Set 1 (New England BioLabs (NEB) E6040, Ipswich, Massachusetts, USA) was combined with the Multiplexing Sample Preparation Oligo Kit (Illumina PE-400-1001). Per sample, 22µl of the prepared sample was subjected to PCR (21 cycles) following the Illumina Library Amplification Index Protocol. After purification (QIAquick PCR Purification columns, Qiagen), elution and concentration (evaporation), the libraries were pooled and denatured (NaOH), followed by Illumina HiSeq 2000 sequencing (multiplexed paired-end run, 2 times 45 cycles) of 10pM/pool.
GEO Accession
GSM2552052