title: GSM1428951: DP Input BGT GLIB-Seq; Mus musculus; MeDIP-Seq
library_construction_protocol: Genomic DNA was prepared by lysing ES or DP T cells, followed by treatment with RNase A (Qiagen) for 1 hour at 37 0C and then overnight treatment with proteinase K (Roche) at 550C with vigorous shaking. DNA was purified after sequential treatment with phenol, phenol/chlorophorm/isoamyl alcohol, chlorophorm/isoamyl alcohol (all obtained from Sigma) and then ethanol (Sigma) precipitation. DNA was resuspended in 10 mM Tris-Cl pH 8.0, 0.1mM EDTA .Biotin-glucosyl tagging and enrichment of 5hmC (Biotin-gmC, a variant of GLIB (Song C.-X. et al., Selective chemical labeling reveals the genome-wide distribution of 5-hydroxymethylcytosine, Nature Biotechnology 29, 68–72 (2011))): For specific biotinylation of 5hmC in DP T cells and ES cells, we used the Hydroxymethyl Collector kit  (Active Motif, Carlsbad, CA) to selectively label 5hmC bases with glucose and biotin moieties to form biotin-N3-5-gmC as previously described(Song C.-X. et al., Selective chemical labeling reveals the genome-wide distribution of 5-hydroxymethylcytosine, Nature Biotechnology 29, 68–72 (2011)). To ensure efficient biotinylation, 2 µg of genomic DNA were biotinylated per reaction. In this case we used the 5500 SoliD fragment library preparation kit from Applied Biosystems in order to prepare the library and the 5500 SOLiD Library indexed adaptors 1-16 according to the manufacturer’s instructions. The samples were sequenced using the SOLiD 5500 platform.