GSM2730268: WT_M0 (5hmC); Mus musculus; OTHER
Genomic DNA was extracted by digesting cells with proteinase K, removal of protein by phenol/chloroform and ethanol precipitation. Genomic DNA was sonicated to ~200bp fragments. Ten micrograms of sonicated genomic DNA were then used to ligate with the adaptor, provided in the NEBNext DNA Library Prep Master Mix Set for Illumina kit (E6040), following the manufacturer's instructions. Sonicated and adaptor-ligated genomic DNA were incubated with 5 μl of 5hmC antibody (Active Motif, 39791) at 4°C overnight in a final volume of 500 μl in DIP buffer (10 mM sodium phosphate, pH 7.0, 140 mM NaCl, 0.05% Triton X-100). The immunoprecipitated DNA fragments were enriched with Dynabeads Protein G (Life Technologies) and amplified with the primers provided in the NEBNext Multiplex Oligos for Illumina preparation kit (E7335). The prepared libraries were sequenced on an Illumina HiSeq2000 machine at Yale Stem Cell Center.
GEO Accession
GSM2730268