GSM4982857: cow_CC_H3K4me3_CR_rep2; Bos taurus; OTHER
CUT&RUN library preparation and sequencing  CUT&RUN was performed as previously reported (Skene et al., 2018; Skene and Henikoff, 2017) with modifications. Briefly, after removing the zona pellucida with Tyrode's solution (Sigma-Aldrich, T1788) or 0.5% (m/v) Protease from Streptomyces griseus (Sigma-Aldrich, P8811), mammalian oocytes and early embryos were incubated with Concanavalin-coated magnetic beads (Polyscience, 86057) for 10 min at room temperature on a thermomixer at 400 rpm. Samples were then incubated with primary antibody at a ratio of 1:100 at 4℃ overnight on a thermomixer at 400 rpm. The next day, after washing for one time, beads were incubated with pA-MNase (to a final concentration of 400-700ng/mL) (a gift from Steven Henikoff lab) at 4℃ for 3 hours on a thermomixer at 400 rpm. After two times of washing, targeted digestion was performed by adding 2μL of 100mM CaCl2 for 30 mins on ice, followed by termination by adding an equal volume of 2  × stop buffer. Samples were then incubated at 37℃ for 20 mins for fragment releasing. The total samples or supernatants were digested with Proteinase K (NEB, P8107S) and purified using phenol:chloroform:isoamyl alcohol (25:24:1, v/v) followed by ethanol purification at -80℃ overnight. The next day, DNA was purified and subjected to Truseq library preparation using NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB, E7645S). Sequencing was done using the HiSeq X Ten system (Illumina) according to the manufacturer's protocol.