GSM5035587: C13-1; Bos taurus; ChIP-Seq
5 µg of the genomic DNA extracted from sperms were fragmented with an S2 ultrasonicator (Covaris, Woburn, MA, USA) in 10 mM Tris buffer (pH 8.0) to obtain approximately 150-bp fragments. After sonication, the fragmented DNA was concentrated to 20 µL in 100 mM NaPi buffer (pH 8.0) using QIAquick PCR purification kit (Qiagen). A 100 µL volume of 100 mM NaPi buffer with 20 mM amine-PEG2-biotin (Thermo Fisher Scientific) was added and the mixture was heated to 75 °C for 10 min. After thermal equilibration to room temperature, 5 mM K2IrBr6 (Sigma-Aldrich) was added for 8-oxoG biotinylation. The biotinylated DNA fragments were eluted with 125 µL Tris buffer using the QIAquick PCR purification kit and collected using Dynabeads MyOne Streptavidin C1 (Thermo Fisher Scientific). Complementary DNA strands to those with bound biotinylated 8-oxoG were released by incubation in 150 mM NaOH at 20 °C for 30 min, and concentrated to 10 µL ddH2O using ssDNA/RNA Clean & Concentrator kit (Zymo Research, Irvine, CA, USA). The OG-seq library was constructed using the TruSeq Nano DNA kit (Illumina, San Diego, CA, USA) ChIP-Seq using captured 8-oxoG containing DNA fragments
GEO Accession
GSM5035587