GSM8447051: D-9_SN_scBS_seq; Mus musculus; Bisulfite-Seq
Fully-grown germinal vesicle (GV) oocytes were mechanically dissected from the ovaries of two 12-week-old C57Bl6/Babr mice in M2 medium (Sigma-Aldrich, M7167). After removing all cumulus cells, oocytes were placed in M2 with Hoechst (5µM, Abcam, 33343) and incubated for 10 minutes at room temperature. Oocytes were washed in PBS and NSN/SN stage was scored on a Zeiss LSM780 confocal microscope. Absence of a ring around the nucleolus was counted as “NSN”, a partial ring as “intermediate” and a full ring “SN”. After scoring, oocytes were frozen individually in 5μl RLT Plus buffer (Qiagen) and stored at -80°C until further use. Single-cell BS-seq (scBS-seq) libraries were prepared as previously described (Smallwood et al., 2014), with minor changes as described in (Castillo-Fernandez et.al, 2020)