GSM1645674: IUGR fetus  in twins 4; Homo sapiens; Bisulfite-Seq
Genomic DNA was extracted using the QIAamp DNA Blood Mini Kit (Qiagen). 1µg of genomic DNA was fragmented into approximately 200-300bp using a Covarias sonication system (Covarias). After purification, the fragmented fractions were treated with a mix of T4 DNA polymerase, Klenow Fragment and T4 polynucleotide kinase to repair blunt and phosphorylate ends. Total RNA was extracted using the Invitrogen TRIzol® Reagent and then treated with RNase-free DNase I (Ambion) for 30 min. The integrity of total RNA was checked using an Agilent 2100 Bioanalyser. After purification, the fragmented fractions were treated with a mix of T4 DNA polymerase, Klenow Fragment and T4 polynucleotide kinase to repair blunt and phosphorylate ends. The blunt DNA fragments were subsequently 3'-adenylated using Klenow Fragment (3-5' exo-) and then ligated by T4 DNA Ligase to adapters that were synthesized with 5'-methylcytosine instead of 5'-cytosine and contain index sequences inside. After each step, the reaction products were purified using QIAquick PCR purification kit (Qiagen).  The constructed libraries were quantified using a Qubit fluorometer with Quant-iT dsDNA HS Assay Kit (Invitrogen).