GSM8247145: rupture_nohCG_rep4; Mus musculus; RNA-Seq
Mouse secondary follicles were isolated from the ovaries of 14-16 day-old CD-1 female mice enzymatically using L15 media (Thermo Fisher Scientific, Waltham, MA) with 1% penicillin-streptomycin (Thermo Fisher Scientific), 25 μg/ml liberase (Sigma-Aldrich, St. Louis, MO), and 200 μg/mlDNase I (Qiagen, Hilden, Germany). Ovary pieces were manually dispersed using a P1000 pipette to release follicles. Follicles with intact morphology were encapsulated in an alginate hydrogel (Sigma-Aldrich) for the preservation of the 3D structure of follicles throughout folliculogenesis. Follicles were cultured for 8 days, then antral follicles were removed, washed, and transferred to maturation media with or without human chorionic gonadotropin (Sigma-Aldrich for 12 hours to allow for ovulation and follicle rupture. Bulk RNA-seq libraries were generated using SMART-Seq2 protocol (Trombetta et al., 2014). Briefly, cDNA was reversed transcribed from single cells using Maxima RT (Thermo Fisher Scientific) and whole transcriptome amplification (WTA) was performed. WTA products were purified using the Agencourt AMPure XP beads (Beckman Coulter, IN, USA) and used to prepare paired-end libraries with Nextera XT  (Illumina, CA, USA).